Inhibitors of cellular necrosis and related methods

ABSTRACT

A compound having the following structure (I): 
     
       
         
         
             
             
         
       
     
     or a pharmaceutically acceptable salt, prodrug, stereoisomer or tautomer thereof, is provided. Related compounds, methods for preparation of the same and uses of the compounds for treatment of various indications, including treatment of necrotic cell diseases and/or inflammation, are also provided.

BACKGROUND

1. Technical Field

This invention generally relates to inhibitors of necrosis and methodsfor their use and preparation. The compounds, and compositionscomprising the same, can be used in methods for preventing and/ortreating diseases involving cell death and/or inflammation.

2. Description of the Related Art

Programmed necrotic cell death, also called necroptosis, is a form ofcell death in which various stimuli such as TNFα, certain toll-likereceptor (TLR) agonists, and ischemia can induce cellular necrosis.Necroptosis is a highly inflammatory form of cell death and is thoughtto be an important contributor to pathology in multiple degenerative andinflammatory diseases. These diseases include neurodegenerativediseases, stroke, coronary heart disease and myocardial infarction,retinal degenerative diseases, inflammatory bowel disease, kidneydisease, liver disease, and others.

Necrosis is characterized by cell membrane and organelle disruption,cell swelling and mitochondrial impairment, followed by cell lysis(Syntichaki, P.; Tavernarakis, N. EMBO Rep. 2002, 3(7), 604-609; Martin,L. J., Al-Abdulla, N. A.; Brambrink, A. M.; Kirsch, J. R.; Sieber, F.E.; Portera-Cailliau, C. Brain Res. Bull. 1998, 46(4), 281-309). Also,cell lyses typically are accompanied by an inflammatory response. Someof the underlying biochemical events in this process are now understood,and the activity of receptor interacting protein kinase 1 (RIP1 kinase)has been shown to be important for cells to undergo necroptosis.Furthermore, RIP1 kinase activity is also known to promote the releaseof inflammatory mediators such as TNF alpha from cells which can induceinflammation and also promote further necroptosis (Christofferson, D.E., Li, Y., Hitomi, J., Zhou, W., Upperman, C., Zhu, H., Gerber, S. A.,Gygi, S., Yuan, J. Cell Death Dis. 2012, 3, e320). Therefore,identifying and preparing low molecular weight molecules that preventnecrotic cell death and/or inflammation by inhibiting RIP1 kinase or byother mechanisms can provide useful compounds for therapeuticintervention in diseases characterized by necrotic cell death and/orinflammation.

Small molecules inhibitors of cellular necrosis have been investigated.For example, U.S. Pat. No. 7,491,743 (“the '743 patent”) and U.S. PatentPublication No. 2012/012,889 describe indole-substituted hydantoinmolecules as inhibitors of necrosis. The compounds disclosed in thesepublications include chiral hydantoin moieties linked to an indolemoiety via a methylene bridge. Although various preparations for suchcompounds have been proposed, each of these preparations suffer fromvarious disadvantages such as achiral products, linear syntheticstrategies, long and/or low yielding synthetic routes and/or use ofenzymes for resolution of racemic products or reagents. Accordingly,none of the currently available preparation methods are useful for largescale production of the indole-substituted hydantoin compounds.

While progress has been made, there remains a need in the art forimproved methods for preparation of inhibitors of cellular necrosis aswell as improved compounds for preventing and treating diseasesinvolving cell death and/or inflammation. The present disclosureprovides this and related benefits.

BRIEF SUMMARY

Embodiments of the present invention provide compounds which areinhibitors of cellular necrosis. Accordingly, the provided compoundsfind utility as therapeutics for treatment of various disordersassociated with cellular necrosis, such as trauma, ischemia, stroke,myocardial infarction, infection, sepsis, Parkinson's disease,Alzheimer's disease, amyotrophic lateral sclerosis, Huntington'sdisease, HIV-associated dementia, retinal degenerative disease,glaucoma, age-related macular degeneration, rheumatoid arthritis,psoriasis, inflammatory bowel disease, kidney disease and others. Thepresent inventors have surprisingly found that compounds of certainembodiments of the invention are more potent inhibitors of cellularnecrosis relative to structurally related compounds, including thosecompounds disclosed in the '743 patent.

Accordingly, in one embodiment, there is provided a compound having thefollowing structure (I):

or a pharmaceutically acceptable salt, prodrug, stereoisomer or tautomerthereof. Pharmaceutical compositions comprising a compound of structure(I), or a pharmaceutically acceptable salt, prodrug, stereoisomer ortautomer thereof, and a pharmaceutically acceptable carrier, diluent orexcipient are also provided.

In yet other embodiments, the disclosure is directed to compounds havingthe following structure (II):

or a pharmaceutically acceptable salt, prodrug, stereoisomer or tautomerthereof, wherein R⁷, R⁸, R⁹ and R¹⁰ are as defined herein. Compositionscomprising compounds of structure (II) or a pharmaceutically acceptablesalt, prodrug or tautomer thereof, and a pharmaceutically acceptablecarrier, diluent or excipient are also provided in various otherembodiments.

In different embodiments, the present invention provides a compound ofstructure (I) or (II) for use as a medicament. For example, in someembodiments the invention is directed to a method for treating anecrotic cell disease, the method comprising administering an effectiveamount of a pharmaceutical composition comprising a compound ofstructure (I) or (II), or a pharmaceutically acceptable salt, prodrug,stereoisomer or tautomer thereof, to a subject in need thereof. Somedifferent embodiments include use of a compound of structure (I) or (II)for treatment of a necrotic cell disease. Other embodiments include useof a compound of structure (I) or (II) for manufacture of a medicamentfor treating a necrotic cell disease.

Exemplary necrotic cell diseases which can be treated by the disclosedmethods include, but are not limited to, trauma, ischemia, stroke,cardiac infarction, infection, Gaucher's disease, Krabbe disease,sepsis, Parkinson's disease, Alzheimer's disease, amyotrophic lateralsclerosis, Huntington's disease, HIV-associated dementia, retinaldegenerative disease, glaucoma, age-related macular degeneration,rheumatoid arthritis, psoriasis, psoriatic arthritis and inflammatorybowel disease.

Still other embodiments are directed to a method for treating aninflammatory disorder, the method comprising administering an effectiveamount of a pharmaceutical composition comprising a compound ofstructure (I) or (II), or a pharmaceutically acceptable salt, prodrug,stereoisomer or tautomer thereof, to a subject in need thereof. Somedifferent embodiments are directed to use of a compound of structure (I)or (II) for treatment of an inflammatory disorder. Other differentembodiments include use of a compound of structure (I) or (II) formanufacture of a medicament for treating an inflammatory disorder.Exemplary inflammatory disorders include, but are not limited toinflammatory bowel disease.

Embodiments of the present invention provide methods for preparation ofhydantoin substituted indole compounds, embodiments of which areinhibitors of cellular necrosis. The provided methods are efficient andamendable to large scale manufacturing of the compounds, as well assmaller scale production for research purposes. Embodiments of thecompounds, which can be prepared according to the disclosed method, findutility as therapeutics for treatment of various disorders associatedwith cellular necrosis, such as trauma, ischemia, stroke, myocardialinfarction, infection, sepsis, Parkinson's disease, Alzheimer's disease,amyotrophic lateral sclerosis, Huntington's disease, HIV-associateddementia, retinal degenerative disease, inflammatory bowel disease,kidney disease and others. Accordingly, in one embodiment, there isprovided a method for preparing a compound having the followingstructure (III):

or a pharmaceutically acceptable salt, prodrug, stereoisomer or tautomerthereof, wherein R¹, R², R³, R⁵, R⁶ are as defined herein, the methodcomprising reacting aldehyde (5), or a salt, stereoisomer or tautomerthereof, with phenyl hydrazine (6), or a salt thereof, to yield (7), ora salt, stereoisomer or tautomer thereof, as follows:

wherein each P is as defined herein.

In other embodiments, the disclosure provides a compound useful forpreparation of the indole-substituted hydantoin compounds (e.g.,compounds of structure (III)), the compound having one of the followingstructures (4′) or (4″):

wherein R′ and P is as defined herein.

These and other aspects of the invention will be apparent upon referenceto the following detailed description. To this end, various referencesare set forth herein which describe in more detail certain backgroundinformation, procedures, compounds and/or compositions, and are eachhereby incorporated by reference in their entirety.

BRIEF DESCRIPTION OF THE DRAWINGS

In the figures, identical reference numbers identify similar elements.The sizes and relative positions of elements in the figures are notnecessarily drawn to scale and some of these elements are arbitrarilyenlarged and positioned to improve figure legibility. Further, theparticular shapes of the elements as drawn are not intended to conveyany information regarding the actual shape of the particular elements,and have been solely selected for ease of recognition in the figures.

FIG. 1 provides RIP1 kinase inhibitory data for a representativecompound and a comparative compound.

DETAILED DESCRIPTION

In the following description, certain specific details are set forth inorder to provide a thorough understanding of various embodiments.However, one skilled in the art will understand that the invention maybe practiced without these details. In other instances, well-knownstructures have not been shown or described in detail to avoidunnecessarily obscuring descriptions of the embodiments. Unless thecontext requires otherwise, throughout the specification and claimswhich follow, the word “comprise” and variations thereof, such as,“comprises” and “comprising” are to be construed in an open, inclusivesense, that is, as “including, but not limited to.” Further, headingsprovided herein are for convenience only and do not interpret the scopeor meaning of the claimed invention.

Reference throughout this specification to “one embodiment” or “anembodiment” means that a particular feature, structure or characteristicdescribed in connection with the embodiment is included in at least oneembodiment. Thus, the appearances of the phrases “in one embodiment” or“in an embodiment” in various places throughout this specification arenot necessarily all referring to the same embodiment. Furthermore, theparticular features, structures, or characteristics may be combined inany suitable manner in one or more embodiments. Also, as used in thisspecification and the appended claims, the singular forms “a,” “an,” and“the” include plural referents unless the content clearly dictatesotherwise. It should also be noted that the term “or” is generallyemployed in its sense including “and/or” unless the content clearlydictates otherwise.

“Amino” refers to the —NH₂ radical.

“Cyano” refers to the —CN radical.

“Hydroxy” or “hydroxyl” refers to the —OH radical.

“Imino” refers to the ═NH substituent.

“Nitro” refers to the —NO₂ radical.

“Oxo” refers to the ═O substituent.

“Thioxo” refers to the ═S substituent.

“Alkyl” refers to a straight or branched hydrocarbon chain radicalconsisting solely of carbon and hydrogen atoms, which is saturated orunsaturated (i.e., contains one or more double “alkenyl” and/or triplebonds “alkynyl”), having from one to twelve carbon atoms (C₁-C₁₂ alkyl),preferably one to eight carbon atoms (C₁-C₈ alkyl) or one to six carbonatoms (C₁-C₆ alkyl), and which is attached to the rest of the moleculeby a single bond, e.g., methyl, ethyl, n-propyl, 1-methylethyl(iso-propyl), n-butyl, n-pentyl, 1,1-dimethylethyl (t-butyl),3-methylhexyl, 2-methylhexyl, ethenyl, prop-1-enyl, but-1-enyl,pent-1-enyl, penta-1,4-dienyl, ethynyl, propynyl, butynyl, pentynyl,hexynyl, and the like. Unless stated otherwise specifically in thespecification, an alkyl group may be optionally substituted.

“Alkylene” or “alkylene chain” refers to a straight or branched divalenthydrocarbon chain linking the rest of the molecule to a radical group,consisting solely of carbon and hydrogen, which is saturated orunsaturated (i.e., contains one or more double and/or triple bonds), andhaving from one to twelve carbon atoms, e.g., methylene, ethylene,propylene, n-butylene, ethenylene, propenylene, n-butenylene,propynylene, n-butynylene, and the like. The alkylene chain is attachedto the rest of the molecule through a single or double bond and to theradical group through a single or double bond. The points of attachmentof the alkylene chain to the rest of the molecule and to the radicalgroup can be through one carbon or any two carbons within the chain.Unless stated otherwise specifically in the specification, an alkylenechain may be optionally substituted.

“Alkoxy” refers to a radical of the formula —OR_(a) where R_(a) is analkyl radical as defined above containing one to twelve carbon atoms.Unless stated otherwise specifically in the specification, an alkoxygroup may be optionally substituted.

“Alkylamino” refers to a radical of the formula —NHR_(a) or —NR_(a)R_(a)where each R_(a) is, independently, an alkyl radical as defined abovecontaining one to twelve carbon atoms. Unless stated otherwisespecifically in the specification, an alkylamino group may be optionallysubstituted.

“Thioalkyl” refers to a radical of the formula —SR_(a) where R_(a) is analkyl radical as defined above containing one to twelve carbon atoms.Unless stated otherwise specifically in the specification, a thioalkylgroup may be optionally substituted.

“Aryl” refers to a hydrocarbon ring system radical comprising hydrogen,6 to 18 carbon atoms and at least one aromatic ring. For purposes ofthis invention, the aryl radical may be a monocyclic, bicyclic,tricyclic or tetracyclic ring system, which may include fused or bridgedring systems. Aryl radicals include, but are not limited to, arylradicals derived from aceanthrylene, acenaphthylene, acephenanthrylene,anthracene, azulene, benzene, chrysene, fluoranthene, fluorene,as-indacene, s-indacene, indane, indene, naphthalene, phenalene,phenanthrene, pleiadene, pyrene, and triphenylene. Unless statedotherwise specifically in the specification, the term “aryl” or theprefix “ar-” (such as in “aralkyl”) is meant to include aryl radicalsthat are optionally substituted.

“Aralkyl” refers to a radical of the formula —R_(b)—R_(c) where R_(b) isan alkylene chain as defined above and R_(c) is one or more arylradicals as defined above, for example, benzyl, diphenylmethyl and thelike. Unless stated otherwise specifically in the specification, anaralkyl group may be optionally substituted.

“Cycloalkyl” or “carbocyclic ring” refers to a stable non-aromaticmonocyclic or polycyclic hydrocarbon radical consisting solely of carbonand hydrogen atoms, which may include fused or bridged ring systems,having from three to fifteen carbon atoms, preferably having from threeto ten carbon atoms, and which is saturated or unsaturated and attachedto the rest of the molecule by a single bond. Monocyclic radicalsinclude, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,cycloheptyl, and cyclooctyl. Polycyclic radicals include, for example,adamantyl, norbornyl, decalinyl, 7,7-dimethyl-bicyclo[2.2.1]heptanyl,and the like. Unless otherwise stated specifically in the specification,a cycloalkyl group may be optionally substituted.

“Cycloalkylalkyl” refers to a radical of the formula —R_(b)R_(d) whereR_(b) is an alkylene chain as defined above and R_(d) is a cycloalkylradical as defined above. Unless stated otherwise specifically in thespecification, a cycloalkylalkyl group may be optionally substituted.

“Fused” refers to any ring structure described herein which is fused toan existing ring structure in the compounds of the invention. When thefused ring is a heterocyclyl ring or a heteroaryl ring, any carbon atomon the existing ring structure which becomes part of the fusedheterocyclyl ring or the fused heteroaryl ring may be replaced with anitrogen atom.

“Halo” or “halogen” refers to bromo, chloro, fluoro or iodo.

“Haloalkyl” refers to an alkyl radical, as defined above, that issubstituted by one or more halo radicals, as defined above, e.g.,trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl,1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, and thelike. Unless stated otherwise specifically in the specification, ahaloalkyl group may be optionally substituted.

“Heterocyclyl” or “heterocyclic ring” refers to a stable 3- to18-membered aromatic or non-aromatic ring radical which consists of twoto twelve carbon atoms and from one to six heteroatoms selected from thegroup consisting of nitrogen, oxygen and sulfur. Heterocycles includealiphatic heterocycles and aromatic heterocycles (heteroaryls). Unlessstated otherwise specifically in the specification, the heterocyclylradical may be a monocyclic, bicyclic, tricyclic or tetracyclic ringsystem, which may include fused or bridged ring systems; and thenitrogen, carbon or sulfur atoms in the heterocyclyl radical may beoptionally oxidized; the nitrogen atom may be optionally quaternized;and the heterocyclyl radical may be partially or fully saturated.Examples of such heterocyclyl radicals include, but are not limited to,dioxolanyl, thienyl[1,3]dithianyl, decahydroisoquinolyl, imidazolinyl,imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl,octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl,2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl,piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl,thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl,thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, and1,1-dioxo-thiomorpholinyl. Unless stated otherwise specifically in thespecification, Unless stated otherwise specifically in thespecification, a heterocyclyl group may be optionally substituted.

“N-heterocyclyl” refers to a heterocyclyl radical as defined abovecontaining at least one nitrogen and where the point of attachment ofthe heterocyclyl radical to the rest of the molecule is through anitrogen atom in the heterocyclyl radical. Unless stated otherwisespecifically in the specification, a N-heterocyclyl group may beoptionally substituted.

“Heterocyclylalkyl” refers to a radical of the formula —R_(b)R_(e) whereR_(b) is an alkylene chain as defined above and R_(e) is a heterocyclylradical as defined above, and if the heterocyclyl is anitrogen-containing heterocyclyl, the heterocyclyl may be attached tothe alkyl radical at the nitrogen atom. Unless stated otherwisespecifically in the specification, a heterocyclylalkyl group may beoptionally substituted.

“Heteroaryl” refers to a 5- to 14-membered ring system radicalcomprising hydrogen atoms, one to thirteen carbon atoms, one to sixheteroatoms selected from the group consisting of nitrogen, oxygen andsulfur, and at least one aromatic ring. For purposes of this invention,the heteroaryl radical may be a monocyclic, bicyclic, tricyclic ortetracyclic ring system, which may include fused or bridged ringsystems; and the nitrogen, carbon or sulfur atoms in the heteroarylradical may be optionally oxidized; the nitrogen atom may be optionallyquaternized. Examples include, but are not limited to, azepinyl,acridinyl, benzimidazolyl, benzothiazolyl, benzindolyl, benzodioxolyl,benzofuranyl, benzooxazolyl, benzothiazolyl, benzothiadiazolyl,benzo[b][1,4]dioxepinyl, 1,4-benzodioxanyl, benzonaphthofuranyl,benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl,benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl(benzothiophenyl), benzotriazolyl, benzo[4,6]imidazo[1,2-a]pyridinyl,carbazolyl, cinnolinyl, dibenzofuranyl, dibenzothiophenyl, furanyl,furanonyl, isothiazolyl, imidazolyl, indazolyl, indolyl, indazolyl,isoindolyl, indolinyl, isoindolinyl, isoquinolyl, indolizinyl,isoxazolyl, naphthyridinyl, oxadiazolyl, 2-oxoazepinyl, oxazolyl,oxiranyl, 1-oxidopyridinyl, 1-oxidopyrimidinyl, 1-oxidopyrazinyl,1-oxidopyridazinyl, 1-phenyl-1H-pyrrolyl, phenazinyl, phenothiazinyl,phenoxazinyl, phthalazinyl, pteridinyl, purinyl, pyrrolyl, pyrazolyl,pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, quinazolinyl,quinoxalinyl, quinolinyl, quinuclidinyl, isoquinolinyl,tetrahydroquinolinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl,triazinyl, and thiophenyl (i.e. thienyl). Unless stated otherwisespecifically in the specification, a heteroaryl group may be optionallysubstituted.

“N-heteroaryl” refers to a heteroaryl radical as defined abovecontaining at least one nitrogen and where the point of attachment ofthe heteroaryl radical to the rest of the molecule is through a nitrogenatom in the heteroaryl radical. Unless stated otherwise specifically inthe specification, an N-heteroaryl group may be optionally substituted.

“Heteroarylalkyl” refers to a radical of the formula —R_(b)R_(f) whereR_(b) is an alkylene chain as defined above and R_(f) is a heteroarylradical as defined above. Unless stated otherwise specifically in thespecification, a heteroarylalkyl group may be optionally substituted.

The term “substituted” used herein means any of the above groups (i.e.,alkyl, alkylene, alkoxy, alkylamino, thioalkyl, aryl, aralkyl,cycloalkyl, cycloalkylalkyl, haloalkyl, heterocyclyl, N-heterocyclyl,heterocyclylalkyl, heteroaryl, N-heteroaryl and/or heteroarylalkyl)wherein at least one hydrogen atom is replaced by a bond to anon-hydrogen atoms such as, but not limited to: an alkyl group, ahalogen atom such as F, Cl, Br, and I; an oxygen atom in groups such ashydroxyl groups, alkoxy groups, and ester groups; a sulfur atom ingroups such as thiol groups, thioalkyl groups, sulfone groups, sulfonylgroups, and sulfoxide groups; a nitrogen atom in groups such as amines,amides, alkylamines, dialkylamines, arylamines, alkylarylamines,diarylamines, N-oxides, imides, and enamines; a silicon atom in groupssuch as trialkylsilyl groups, dialkylarylsilyl groups, alkyldiarylsilylgroups, and triarylsilyl groups; and other heteroatoms in various othergroups. “Substituted” also means any of the above groups in which one ormore hydrogen atoms are replaced by a higher-order bond (e.g., a double-or triple-bond) to a heteroatom such as oxygen in oxo, carbonyl,carboxyl, and ester groups; and nitrogen in groups such as imines,oximes, hydrazones, and nitriles. For example, “substituted” includesany of the above groups in which one or more hydrogen atoms are replacedwith —NR_(g)R_(h), —NR_(g)C(═O)R_(h), —NR_(g)C(═O)NR_(g)R_(h),—NR_(g)C(═O)OR_(h), —NR_(g)SO₂R_(h), —OC(═O)NR_(g)R_(h), —OR_(g),—SR_(g), —SOR_(g), —SO₂R_(g), —OSO₂R_(g), —SO₂OR_(g), ═NSO₂R_(g), and—SO₂NR_(g)R_(h). “Substituted” also means any of the above groups inwhich one or more hydrogen atoms are replaced with —C(═O)R_(g),—C(═O)OR_(g), —C(═O)NR_(g)R_(h), —CH₂SO₂R_(g), —CH₂SO₂NR_(g)R_(h). Inthe foregoing, R_(g) and R_(h) are the same or different andindependently hydrogen, alkyl, alkoxy, alkylamino, thioalkyl, aryl,aralkyl, cycloalkyl, cycloalkylalkyl, haloalkyl, heterocyclyl,N-heterocyclyl, heterocyclylalkyl, heteroaryl, N-heteroaryl and/orheteroarylalkyl. “Substituted” further means any of the above groups inwhich one or more hydrogen atoms are replaced by a bond to an amino,cyano, hydroxyl, imino, nitro, oxo, thioxo, halo, alkyl, alkoxy,alkylamino, thioalkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl,haloalkyl, heterocyclyl, N-heterocyclyl, heterocyclylalkyl, heteroaryl,N-heteroaryl and/or heteroarylalkyl group. In addition, each of theforegoing substituents may also be optionally substituted with one ormore of the above substituents.

The term “leaving group” refers to a molecular fragment that departswith a pair of electrons in heterolytic bond cleavage. Leaving groupscan be charged or neutral molecules. “Leaving groups” include, but arenot limited to, halides such as Cl⁻, Br⁻, sulfonate esters (e.g., TsO⁻),water and ammonia.

“Prodrug” is meant to indicate a compound that may be converted underphysiological conditions or by solvolysis to a biologically activecompound of the invention. Thus, the term “prodrug” refers to ametabolic precursor of a compound of the invention that ispharmaceutically acceptable. A prodrug may be inactive when administeredto a subject in need thereof, but is converted in vivo to an activecompound of the invention. Prodrugs are typically rapidly transformed invivo to yield the parent compound of the invention, for example, byhydrolysis in blood. The prodrug compound often offers advantages ofsolubility, tissue compatibility or delayed release in a mammalianorganism (see, Bundgard, H., Design of Prodrugs (1985), pp. 7-9, 21-24(Elsevier, Amsterdam)). A discussion of prodrugs is provided in Higuchi,T., et al., A.C.S. Symposium Series, Vol. 14, and in BioreversibleCarriers in Drug Design, Ed. Edward B. Roche, American PharmaceuticalAssociation and Pergamon Press, 1987.

The term “prodrug” is also meant to include any covalently bondedcarriers, which release the active compound of the invention in vivowhen such prodrug is administered to a mammalian subject. Prodrugs of acompound of the invention may be prepared by modifying functional groupspresent in the compound of the invention in such a way that themodifications are cleaved, either in routine manipulation or in vivo, tothe parent compound of the invention. Prodrugs include compounds of theinvention wherein a hydroxy, amino or mercapto group is bonded to anygroup that, when the prodrug of the compound of the invention isadministered to a mammalian subject, cleaves to form a free hydroxy,free amino or free mercapto group, respectively. Examples of prodrugsinclude, but are not limited to, acetate, formate and benzoatederivatives of alcohol or amide derivatives of amine functional groupsin the compounds of the invention and the like.

The invention disclosed herein is also meant to encompass allpharmaceutically acceptable compounds of structure (I) beingisotopically-labelled by having one or more atoms replaced by an atomhaving a different atomic mass or mass number. Examples of isotopes thatcan be incorporated into the disclosed compounds include isotopes ofhydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine, chlorine, andiodine, such as ²H, ³H, ¹¹C, ¹³C, ¹⁴C, ¹³N, ¹⁵N, ¹⁵O, ¹⁷O, ¹⁸O, ³¹P,³²P, ³⁵S, ³⁶Cl, ¹²³I, and ¹²⁵I, respectively. These radiolabeledcompounds could be useful to help determine or measure the effectivenessof the compounds, by characterizing, for example, the site or mode ofaction, or binding affinity to pharmacologically important site ofaction. Certain isotopically-labelled compounds of structure (I), forexample, those incorporating a radioactive isotope, are useful in drugand/or substrate tissue distribution studies. The radioactive isotopestritium, i.e. ³H, and carbon-14, i.e. ¹⁴C, are particularly useful forthis purpose in view of their ease of incorporation and ready means ofdetection.

Substitution with heavier isotopes such as deuterium, i.e. ²H, mayafford certain therapeutic advantages resulting from greater metabolicstability, for example, increased in vivo half-life or reduced dosagerequirements, and hence may be preferred in some circumstances.Substitution with positron emitting isotopes, such as ¹¹C, ¹⁸F, ¹⁵C and¹³N, can be useful in Positron Emission Topography (PET) studies forexamining substrate receptor occupancy. Isotopically-labeled compoundsof structure (I) can generally be prepared by conventional techniquesknown to those skilled in the art or by processes analogous to thosedescribed in the Preparations and Examples as set out below using anappropriate isotopically-labeled reagent in place of the non-labeledreagent previously employed.

The invention disclosed herein is also meant to encompass the in vivometabolic products of the disclosed compounds. Such products may resultfrom, for example, the oxidation, reduction, hydrolysis, amidation,esterification, and the like of the administered compound, primarily dueto enzymatic processes. Accordingly, the invention includes compoundsproduced by a process comprising administering a compound of thisinvention to a mammal for a period of time sufficient to yield ametabolic product thereof. Such products are typically identified byadministering a radiolabeled compound of the invention in a detectabledose to an animal, such as rat, mouse, guinea pig, monkey, or to human,allowing sufficient time for metabolism to occur, and isolating itsconversion products from the urine, blood or other biological samples.

“Stable compound” and “stable structure” are meant to indicate acompound that is sufficiently robust to survive isolation to a usefuldegree of purity from a reaction mixture, and formulation into anefficacious therapeutic agent.

“Mammal” includes humans and both domestic animals such as laboratoryanimals and household pets (e.g., cats, dogs, swine, cattle, sheep,goats, horses, rabbits), and non-domestic animals such as wildlife andthe like.

Often crystallizations produce a solvate of the compound of theinvention. As used herein, the term “solvate” refers to an aggregatethat comprises one or more molecules of a compound of the invention withone or more molecules of solvent. The solvent may be water, in whichcase the solvate may be a hydrate. Alternatively, the solvent may be anorganic solvent. Thus, the compounds of the present invention may existas a hydrate, including a monohydrate, dihydrate, hemihydrate,sesquihydrate, trihydrate, tetrahydrate and the like, as well as thecorresponding solvated forms. The compound of the invention may be truesolvates, while in other cases, the compound of the invention may merelyretain adventitious water or be a mixture of water plus someadventitious solvent.

“Treating” or “treatment” as used herein covers the treatment of thedisease or condition of interest in a mammal (e.g., conditionsassociated with cellular necrosis), preferably a human, having thedisease or condition of interest, and includes:

(i) preventing the disease or condition from occurring in a mammal, inparticular, when such mammal is predisposed to the condition but has notyet been diagnosed as having it;

(ii) inhibiting the disease or condition, i.e., arresting itsdevelopment;

(iii) relieving the disease or condition, i.e., causing regression ofthe disease or condition; or

(iv) relieving the symptoms resulting from the disease or condition,i.e., relieving pain without addressing the underlying disease orcondition. As used herein, the terms “disease” and “condition” may beused interchangeably or may be different in that the particular maladyor condition may not have a known causative agent (so that etiology hasnot yet been worked out) and it is therefore not yet recognized as adisease but only as an undesirable condition or syndrome, wherein a moreor less specific set of symptoms have been identified by clinicians.

The compounds of the invention, or their pharmaceutically acceptablesalts include an asymmetric center and may thus give rise toenantiomers, diastereomers, and other stereoisomeric forms that may bedefined, in terms of absolute stereochemistry, as (R)- or (S)- or, as(D)- or (L)- for amino acids. The present invention is meant to includeall such possible isomers, as well as their racemic and optically pureforms. Optically active (+) and (−), (R)- and (S)-, or (D)- and(L)-isomers may be prepared using chiral synthons or chiral reagents, orresolved using conventional techniques, for example, chromatography andfractional crystallization. Conventional techniques for thepreparation/isolation of individual enantiomers include chiral synthesisfrom a suitable optically pure precursor or resolution of the racemate(or the racemate of a salt or derivative) using, for example, chiralhigh pressure liquid chromatography (HPLC). When the compounds describedherein contain olefinic double bonds or other centres of geometricasymmetry, and unless specified otherwise, it is intended that thecompounds include both E and Z geometric isomers. Likewise, alltautomeric forms are also intended to be included.

A “stereoisomer” refers to a compound made up of the same atoms bondedby the same bonds but having different three-dimensional structures,which are not interchangeable. The present invention contemplatesvarious stereoisomers and mixtures thereof and includes “enantiomers”,which refers to two stereoisomers whose molecules are nonsuperimposeablemirror images of one another.

A “tautomer” refers to a proton shift from one atom of a molecule toanother atom of the same molecule. The present invention includestautomers of any said compounds.

The term “necrotic cell disease” refers to diseases associated with orcaused by cellular necrosis. Exemplary necrotic cell diseases include,but are not limited to, acute diseases such as trauma, ischemia, stroke,cardiac infarction, anthrax lethal toxin induced septic shock, sepsis,cell death induced by LPS, and HIV induced T-cell death leading toimmunodeficiency. The term “necrotic cell disease” also includes but isnot limited to chronic neurodegenerative diseases, such as Parkinson'sdisease, Huntington's disease, amyotrophic lateral sclerosis,Alzheimer's disease, infectious encelopathies, dementia such as HIVassociated dementia. The term “necrotic cell disease” also includes butis not limited to diseases such as inflammatory bowel disease and acuteand chronic kidney disease which are characterized by inflammation andcell death.

The term “ED₅₀” means the dose of a drug that produces 50% of itsmaximum response or effect. Alternatively, “ED₅₀” means the dose thatproduces a pre-determined response in 50% of test subjects orpreparations.

The term “LD₅₀” means the dose of a drug that is lethal in 50% of testsubjects.

The term “EC₅₀” means the concentration of a drug that produces 50% ofits maximum response or effect in a test assay. Alternatively, “EC₅₀”means the effective concentration that produces a pre-determinedresponse in 50% of test assays.

The term “therapeutic index” refers to the therapeutic index of a drugdefined as LD₅₀/ED₅₀.

The chemical names used herein are generated using the ChemDraw UltraVersion 11.0 software naming program (CambridgeSoft).

I. Compounds

Compounds of Structure (I)

As noted above, certain embodiments of the present disclosure aredirected to a compound having the following structure (I):

or a pharmaceutically acceptable salt, prodrug, stereoisomer or tautomerthereof.

Compounds of structure (I) in enantiomerically enriched orenantiomerically pure form are also provided. Accordingly, in someembodiments the compound has the following structure (Ia):

or a pharmaceutically acceptable salt, prodrug or tautomer thereof.

In other embodiments, the compound has the following structure (Ib):

or a pharmaceutically acceptable salt, prodrug or tautomer thereof.

In some embodiments, compound (I), (Ia) or (Ib) is provided in the formof a pharmaceutically acceptable salt. Exemplary salts for this purposeare described herein.

In some other embodiments, the invention provides a prodrug whichconverts to compound (I), (Ia) or (Ib) in vivo. Exemplary prodrugs forthis purpose are known in the art and described herein.

The following General Reaction Schemes I and II illustrate exemplarymethods of making compounds of structure (I):

Other methods for preparing compounds of structure (I) are providedherein below and in the Examples (e.g., Example 1). It is understoodthat one skilled in the art may be able to make these compounds bysimilar methods or by combining other methods known to one skilled inthe art. In general, starting components and reagents may be obtainedfrom sources such as Sigma Aldrich, Lancaster Synthesis, Inc.,Maybridge, Matrix Scientific, TCl, and Fluorochem USA, etc. orsynthesized according to sources known to those skilled in the art (see,for example, Advanced Organic Chemistry: Reactions, Mechanisms, andStructure, 5th edition (Wiley, December 2000)) or prepared as describedin this invention.

For example, compounds of structure (I) may be prepared with referenceto the following General Reaction Scheme I:

Referring to General Reaction Scheme 1, indole A is treated with POCl₃to obtain aldol B. Reductive alkylation of B and C results in compoundD, which is then reduced to yield compounds of structure (I).

When enantiomerically pure or enriched compounds are desired, thecompounds can be prepared according to General Reaction Scheme II.

Referring to General Reaction Scheme II, appropriately substitutedN-acetyl DL tryptophan E is treated with D-aminoacylase to remove theacetyl group from the D tryptophan moiety. Compounds F and G are thenseparated and treated in parallel reaction pathways. Compound F ismethylated in the presence of thionyl chloride and the resultingcompound is converted to the methyl amide H. Treatment of H withtriphosgene results in compound (Ia).

In a separate reaction pathway, the N-acetyl group of compound G isfirst removed by treatment with HCl. Conditions similar to thosedescribed for conversion of F to (Ia) are then employed to prepare (Ib).It will be apparent to one of ordinary skill in the art that ananalogous method, which employs L-aminoacylase may also be employed.

It will also be appreciated by those skilled in the art that in theprocess described herein the functional groups of intermediate compoundsmay need to be protected by suitable protecting groups. Such functionalgroups include hydroxy, amino, mercapto and carboxylic acid. Suitableprotecting groups for hydroxy include trialkylsilyl or diarylalkylsilyl(for example, t-butyldimethylsilyl, t-butyldiphenylsilyl ortrimethylsilyl), tetrahydropyranyl, benzyl, and the like. Suitableprotecting groups for amino, amidino and guanidino includet-butoxycarbonyl, benzyloxycarbonyl, and the like. Suitable protectinggroups for mercapto include —C(O)—R″ (where R″ is alkyl, aryl orarylalkyl), p-methoxybenzyl, trityl and the like. Suitable protectinggroups for carboxylic acid include alkyl, aryl or arylalkyl esters.Protecting groups may be added or removed in accordance with standardtechniques, which are known to one skilled in the art and as describedherein. The use of protecting groups is described in detail in Green, T.W. and P. G. M. Wutz, Protective Groups in Organic Synthesis (1999), 3rdEd., Wiley. As one of skill in the art would appreciate, the protectinggroup may also be a polymer resin such as a Wang resin, Rink resin or a2-chlorotrityl-chloride resin.

It will also be appreciated by those skilled in the art, although suchprotected derivatives of compounds of this invention may not possesspharmacological activity as such, they may be administered to a mammaland thereafter metabolized in the body to form compounds of theinvention which are pharmacologically active. Such derivatives maytherefore be described as “prodrugs”. All prodrugs of compounds of thisinvention are included within the scope of the invention.

Furthermore, all compounds of structure (I) which exist in free base oracid form can be converted to their pharmaceutically acceptable salts bytreatment with the appropriate inorganic or organic base or acid bymethods known to one skilled in the art. Salts of the compounds of theinvention can be converted to their free base or acid form by standardtechniques.

Compounds of Structure (II)

In other embodiments, the invention is directed to compounds having thefollowing structure (II):

or a pharmaceutically acceptable salt, prodrug, stereoisomer or tautomerthereof, wherein one or two of R⁷, R⁸ or R⁹ is F, and each remaining R⁷,R⁸ or R⁹ is H; and R¹⁰ is H or F.

In some embodiments, the compound has the following structure (IIa):

In some embodiments, the compound has the following structure (IIb):

In some embodiments of structure (II), R¹⁰ H. In other embodiments, R¹⁰F.

In some other embodiments R⁷ is F. In different embodiments R⁸ is F. Inyet other embodiments, R⁹ is F.

In some more specific embodiments, R⁷ and R⁸ are F. In some otherspecific embodiments, R⁸ and R⁹ are F.

In various other embodiments, the compound of structure (II) has one ofthe structures provided in Table 1.

TABLE 1 Exemplary Compounds of structure (II) No. Structure Name II-1

5-((7-chloro-1H-indol-3-yl)methyl)- 3-(2-fluorobenzyl)imidazolidine-2,4-dione II-2

5-((7-chloro-1H-indol-3-yl)methyl)- 3-(3-fluorobenzyl)imidazolidine-2,4-dione II-3

5-((7-chloro-1H-indol-3-yl)methyl)- 3-(3,4-difluorobenzyl)imidazolidine-2,4-dione II-4

5-((7-chloro-1H-indol-3-yl)methyl)- 3-(4-fluorobenzyl)imidazolidine-2,4-dione II-5

5-((7-chloro-6-fluoro-1H-indol-3- yl)methyl)-3-(2-fluorobenzyl)imidazolidine-2,4- dione II-6

5-((7-chloro-6-fluoro-1H-indol-3- yl)methyl)-3-(2,3-difluorobenzyl)imidazolidine-2,4- dione II-7

5-((7-chloro-6-fluoro-1H-indol-3- yl)methyl)-3-(3,4-difluorobenzyl)imidazolidine-2,4- dione

Enantiomerically pure or enriched compounds of the compounds in Table 1,including those exemplified in the Examples which follow, are alsoprovided

As described in more detail in the Examples, the compounds of structure(II), such as those compounds illustrated in Table 1, were found to besurprisingly more potent inhibitors of necrosis relative to astructurally related compound disclosed in U.S. Patent Publication No.2012/0122889, which is hereby incorporated by reference in its entiretywith respect to small-molecule inhibitors of necrosis. Compounds ofstructure (II) may be prepared according to the procedures describedherein below and in the Examples which follow.

II. Compositions

For the purposes of administration, the compounds of the presentinvention may be administered as a raw chemical or may be formulated aspharmaceutical compositions. Pharmaceutical compositions of the presentinvention comprise a compound of structure (I) or (II) and apharmaceutically acceptable carrier, diluent or excipient. Accordingly,different embodiments are directed to pharmaceutical compositionscomprising any one or more of the foregoing compounds (e.g., compoundsof structure (I) or (II)) or a pharmaceutically acceptable salt,prodrug, stereoisomer or tautomer thereof, and a pharmaceuticallyacceptable carrier, diluent or excipient are also provided in variousembodiments.

The pharmaceutical compositions of the present invention may bespecially formulated for administration in solid or liquid form,including those adapted for the following: oral administration, forexample, drenches (aqueous or non-aqueous solutions or suspensions),tablets, e.g., those targeted for buccal, sublingual, and systemicabsorption, boluses, powders, granules, pastes for application to thetongue; parenteral administration, for example, by subcutaneous,intramuscular, intravenous or epidural injection as, for example, asterile solution or suspension, or sustained-release formulation;topical application, for example, as a cream, ointment, or acontrolled-release patch or spray applied to the skin; intravaginally orintrarectally, for example, as a pessary, cream or foam; sublingually;ocularly; transdermally; or nasally, pulmonary and to other mucosalsurfaces.

The phrase “pharmaceutically acceptable” is employed herein to refer tothose compounds, materials, compositions, and/or dosage forms which are,within the scope of sound medical judgment, suitable for use in contactwith the tissues of human beings and animals without excessive toxicity,irritation, allergic response, or other problem or complication,commensurate with a reasonable benefit/risk ratio.

The phrase “pharmaceutically-acceptable carrier” as used herein means apharmaceutically-acceptable material, composition or vehicle, such as aliquid or solid filler, diluent, excipient, or solvent encapsulatingmaterial, involved in carrying or transporting the subject compound fromone organ, or portion of the body, to another organ, or portion of thebody. Each carrier must be “acceptable” in the sense of being compatiblewith the other ingredients of the formulation and not injurious to thepatient. Some examples of materials which can serve aspharmaceutically-acceptable carriers include: sugars, such as lactose,glucose and sucrose; starches, such as corn starch and potato starch;cellulose, and its derivatives, such as sodium carboxymethyl cellulose,ethyl cellulose and cellulose acetate; surfactants, such as polysorbate80 (i.e. Tween 80); powdered tragacanth; malt; gelatin; talc;excipients, such as cocoa butter and suppository waxes; oils, such aspeanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, cornoil and soybean oil; glycols, such as propylene glycol; polyols, such asglycerin, sorbitol, mannitol and polyethylene glycol; esters, such asethyl oleate and ethyl laurate; agar; buffering agents, such asmagnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-freewater; isotonic saline; Ringer's solution; ethyl alcohol; pH bufferedsolutions; polyesters, polycarbonates and/or polyanhydrides; and othernon-toxic compatible substances employed in pharmaceutical formulations.Examples of such formulations include, but are not limited to DMSO, 10mM DMSO, 8% hydroxypropyl-beta-cyclodextrin in PBS, propylene glycol,etc. For example, in a certain embodiment the compounds of the inventioncan be used as 4 mM solution in 8% hydroxypropyl-beta-cyclodextrin inPBS for parenteral administration. In another certain embodiment, thecompounds of the invention can be used as a suspension in 0.5% aqueousCMC containing 0.1% TWEEN 80.

As set out herein, certain embodiments of the present compounds maycontain a basic functional group, such as amino or methylamino (NCH₃),and are, thus, capable of forming pharmaceutically-acceptable salts withpharmaceutically-acceptable acids. The term “pharmaceutically-acceptablesalts” in this respect refers to the relatively non-toxic, inorganic andorganic acid addition salts of compounds of the present invention. Thesesalts can be prepared in situ in the administration vehicle or thedosage form manufacturing process, or by separately reacting a purifiedcompound of the invention in its free base form with a suitable organicor inorganic acid, and isolating the salt thus formed during subsequentpurification. Representative salts include the hydrobromide,hydrochloride, sulfate, bisulfate, phosphate, nitrate, acetate,valerate, oleate, palmitate, stearate, laurate, benzoate, lactate,phosphate, tosylate, citrate, maleate, fumarate, succinate, tartrate,napthylate, mesylate, glucoheptonate, lactobionate, and laurylsulphonatesalts and the like. (See, for example, Berge et al. (1977)“Pharmaceutical Salts”, J. Pharm. Sci. 66:1-19.)

The pharmaceutically acceptable salts of the subject compounds includethe conventional nontoxic salts or quaternary ammonium salts of thecompounds, e.g., from non-toxic organic or inorganic acids. For example,such conventional nontoxic salts include those derived from inorganicacids such as hydrochloride, hydrobromic, sulfuric, sulfamic,phosphoric, nitric, and the like; and the salts prepared from organicacids such as acetic, propionic, succinic, glycolic, stearic, lactic,malic, tartaric, citric, ascorbic, palmitic, maleic, hydroxymaleic,phenylacetic, glutamic, benzoic, salicyclic, sulfanilic,2-acetoxybenzoic, fumaric, toluenesulfonic, methanesulfonic, ethanedisulfonic, oxalic, isothionic, and the like.

In other cases, the compounds of the present invention may contain oneor more acidic functional groups and, thus, are capable of formingpharmaceutically-acceptable salts with pharmaceutically-acceptablebases. The term “pharmaceutically-acceptable salts” in these instancesrefers to the relatively non-toxic, inorganic and organic base additionsalts of compounds of the present invention. These salts can likewise beprepared in situ in the administration vehicle or the dosage formmanufacturing process, or by separately reacting the purified compoundin its free acid form with a suitable base, such as the hydroxide,carbonate or bicarbonate of a pharmaceutically-acceptable metal cation,with ammonia, or with a pharmaceutically-acceptable organic primary,secondary or tertiary amine. Representative alkali or alkaline earthsalts include the lithium, sodium, potassium, calcium, magnesium, andaluminum salts and the like. Representative organic amines useful forthe formation of base addition salts include ethylamine, diethylamine,ethylenediamine, ethanolamine, diethanolamine, piperazine and the like.

Wetting agents, emulsifiers and lubricants, such as sodium laurylsulfate and magnesium stearate, as well as coloring agents, releaseagents, coating agents, sweetening, flavoring and perfuming agents,preservatives and antioxidants can also be present in the compositions.

Examples of pharmaceutically-acceptable antioxidants include: watersoluble antioxidants, such as ascorbic acid, cysteine hydrochloride,sodium bisulfate, sodium metabisulfite, sodium sulfite and the like;oil-soluble antioxidants, such as ascorbyl palmitate, butylatedhydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propylgallate, alpha-tocopherol, and the like; and metal chelating agents,such as citric acid, ethylenediamine tetraacetic acid (EDTA), sorbitol,tartaric acid, phosphoric acid, and the like.

Formulations of the present invention include those suitable for oral,nasal, topical (including buccal and sublingual), rectal, vaginal and/orparenteral administration. The formulations may conveniently bepresented in unit dosage form and may be prepared by any methods wellknown in the art of pharmacy. The amount of active ingredient which canbe combined with a carrier material to produce a single dosage form willvary depending upon the host being treated, the particular mode ofadministration. The amount of active ingredient that can be combinedwith a carrier material to produce a single dosage form will generallybe that amount of the compound which produces a therapeutic effect.Generally, this amount will range from about 1% to about 99% of activeingredient, preferably from about 5% to about 70%, most preferably fromabout 10% to about 30%.

In certain embodiments, a formulation of the present invention comprisesan excipient selected from the group consisting of cyclodextrins,liposomes, micelle forming agents, e.g., bile acids, and polymericcarriers, e.g., polyesters and polyanhydrides; and a compound of thepresent invention. In certain embodiments, an aforementioned formulationrenders orally bioavailable a compound of the present invention.

Methods of preparing these formulations or compositions include the stepof bringing into association a compound of the present invention withthe carrier and, optionally, one or more accessory ingredients. Ingeneral, the formulations are prepared by uniformly and intimatelybringing into association a compound of the present invention withliquid carriers, or finely divided solid carriers, or both, and then, ifnecessary, shaping the product.

Formulations of the invention suitable for oral administration may be inthe form of capsules, cachets, pills, tablets, lozenges (using aflavored basis, usually sucrose and acacia or tragacanth), powders,granules, or as a solution or a suspension in an aqueous or non-aqueousliquid, or as an oil-in-water or water-in-oil liquid emulsion, or as anelixir or syrup, or as pastilles (using an inert base, such as gelatinand glycerin, or sucrose and acacia) and/or as mouth washes and thelike, each containing a predetermined amount of a compound of thepresent invention as an active ingredient. A compound of the presentinvention may also be administered as a bolus, electuary or paste.

In solid dosage forms of the invention for oral administration(capsules, tablets, pills, dragees, powders, granules and the like), theactive ingredient is mixed with one or more pharmaceutically-acceptablecarriers, such as sodium citrate or dicalcium phosphate, and/or any ofthe following: fillers or extenders, such as starches, lactose, sucrose,glucose, mannitol, and/or silicic acid; binders, such as, for example,carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone,sucrose and/or acacia; humectants, such as glycerol; disintegratingagents, such as agar-agar, calcium carbonate, potato or tapioca starch,alginic acid, certain silicates, and sodium carbonate; solutionretarding agents, such as paraffin; absorption accelerators, such asquaternary ammonium compounds; wetting agents, such as, for example,cetyl alcohol, glycerol monostearate, and non-ionic surfactants;absorbents, such as kaolin and bentonite clay; lubricants, such a talc,calcium stearate, magnesium stearate, solid polyethylene glycols, sodiumlauryl sulfate, and mixtures thereof; and coloring agents. In the caseof capsules, tablets and pills, the pharmaceutical compositions may alsocomprise buffering agents. Solid compositions of a similar type may alsobe employed as fillers in soft and hard-shelled gelatin capsules usingsuch excipients as lactose or milk sugars, as well as high molecularweight polyethylene glycols and the like.

A tablet may be made by compression or molding, optionally with one ormore accessory ingredients. Compressed tablets may be prepared usingbinder (for example, gelatin or hydroxypropylmethyl cellulose),lubricant, inert diluent, preservative, disintegrant (for example,sodium starch glycolate or cross-linked sodium carboxymethyl cellulose),surface-active or dispersing agent. Molded tablets may be made in asuitable machine in which a mixture of the powdered compound ismoistened with an inert liquid diluent.

The tablets, and other solid dosage forms of the pharmaceuticalcompositions of the present invention, such as dragees, capsules, pillsand granules, may optionally be scored or prepared with coatings andshells, such as enteric coatings and other coatings well known in thepharmaceutical-formulating art. They may also be formulated so as toprovide slow or controlled release of the active ingredient thereinusing, for example, hydroxypropylmethyl cellulose in varying proportionsto provide the desired release profile, other polymer matrices,liposomes and/or microspheres. They may be formulated for rapid release,e.g., freeze-dried. They may be sterilized by, for example, filtrationthrough a bacteria-retaining filter, or by incorporating sterilizingagents in the form of sterile solid compositions that can be dissolvedin sterile water, or some other sterile injectable medium immediatelybefore use. These compositions may also optionally contain opacifyingagents and may be of a composition that they release the activeingredient(s) only, or preferentially, in a certain portion of thegastrointestinal tract, optionally, in a delayed manner. Examples ofembedding compositions that can be used include polymeric substances andwaxes. The active ingredient can also be in micro-encapsulated form, ifappropriate, with one or more of the above-described excipients.

Liquid dosage forms for oral administration of the compounds of theinvention include pharmaceutically acceptable emulsions, microemulsions,solutions, suspensions, syrups and elixirs. In addition to the activeingredient, the liquid dosage forms may contain inert diluents commonlyused in the art, such as, for example, water or other solvents,solubilizing agents and emulsifiers, such as ethyl alcohol, isopropylalcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzylbenzoate, propylene glycol, 1,3-butylene glycol, oils (in particular,cottonseed, groundnut, corn, germ, olive, castor and sesame oils),glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty acidesters of sorbitan, and mixtures thereof.

Besides inert diluents, the oral compositions can also include adjuvantssuch as wetting agents, emulsifying and suspending agents, sweetening,flavoring, coloring, perfuming and preservative agents.

Suspensions, in addition to the active compounds, may contain suspendingagents as, for example, ethoxylated isostearyl alcohols, polyoxyethylenesorbitol and sorbitan esters, microcrystalline cellulose, aluminummetahydroxide, bentonite, agar-agar and tragacanth, and mixturesthereof.

Formulations of the pharmaceutical compositions of the invention forrectal or vaginal administration may be presented as a suppository,which may be prepared by mixing one or more compounds of the inventionwith one or more suitable nonirritating excipients or carrierscomprising, for example, cocoa butter, polyethylene glycol, asuppository wax or a salicylate, and which is solid at room temperature,but liquid at body temperature and, therefore, will melt in the rectumor vaginal cavity and release the active compound.

Formulations of the present invention which are suitable for vaginaladministration also include pessaries, tampons, creams, gels, pastes,foams or spray formulations containing such carriers as are known in theart to be appropriate.

Dosage forms for the topical or transdermal administration of a compoundof this invention include powders, sprays, ointments, pastes, creams,lotions, gels, solutions, patches and inhalants. The active compound maybe mixed under sterile conditions with a pharmaceutically-acceptablecarrier, and with any preservatives, buffers, or propellants which maybe required.

The ointments, pastes, creams and gels may contain, in addition to anactive compound of this invention, excipients, such as animal andvegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulosederivatives, polyethylene glycols, silicones, bentonites, silicic acid,talc and zinc oxide, or mixtures thereof.

Powders and sprays can contain, in addition to a compound of thisinvention, excipients such as lactose, talc, silicic acid, aluminumhydroxide, calcium silicates and polyamide powder, or mixtures of thesesubstances. Sprays can additionally contain customary propellants, suchas chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons,such as butane and propane.

Transdermal patches have the added advantage of providing controlleddelivery of a compound of the present invention to the body. Dissolvingor dispersing the compound in the proper medium can make such dosageforms. Absorption enhancers can also be used to increase the flux of thecompound across the skin. Either providing a rate controlling membraneor dispersing the compound in a polymer matrix or gel can control therate of such flux.

Ophthalmic formulations, eye ointments, powders, solutions and the like,are also contemplated as being within the scope of this invention.

Pharmaceutical compositions of this invention suitable for parenteraladministration comprise one or more compounds of the invention incombination with one or more pharmaceutically-acceptable sterileisotonic aqueous or nonaqueous solutions, dispersions, suspensions oremulsions, or sterile powders which may be reconstituted into sterileinjectable solutions or dispersions just prior to use, which may containsugars, alcohols, antioxidants, buffers, bacteriostats, solutes whichrender the formulation isotonic with the blood of the intended recipientor suspending or thickening agents.

Examples of suitable aqueous and nonaqueous carriers, which may beemployed in the pharmaceutical compositions of the invention includewater, ethanol, polyols (such as glycerol, propylene glycol,polyethylene glycol, and the like), and suitable mixtures thereof,vegetable oils, such as olive oil, and injectable organic esters, suchas ethyl oleate. Proper fluidity can be maintained, for example, by theuse of coating materials, such as lecithin, by the maintenance of therequired particle size in the case of dispersions, and by the use ofsurfactants.

These compositions may also contain adjuvants such as preservatives,wetting agents, emulsifying agents and dispersing agents. Prevention ofthe action of microorganisms upon the subject compounds may be ensuredby the inclusion of various antibacterial and antifungal agents, forexample, paraben, chlorobutanol, phenyl sorbic acid, and the like. Itmay also be desirable to include isotonic agents, such as sugars, sodiumchloride, and the like into the compositions. In addition, prolongedabsorption of the injectable pharmaceutical form may be brought about bythe inclusion of agents which delay absorption such as aluminummonostearate and gelatin.

In some cases, in order to prolong the effect of a drug, it is desirableto slow the absorption of the drug from subcutaneous or intramuscularinjection. This may be accomplished by the use of a liquid suspension ofcrystalline or amorphous material having poor water solubility. The rateof absorption of the drug then depends upon its rate of dissolution,which in turn, may depend upon crystal size and crystalline form.Alternatively, delayed absorption of a parenterally-administered drugform is accomplished by dissolving or suspending the drug in an oilvehicle.

Injectable depot forms are made by forming microencapsule matrices ofthe subject compounds in biodegradable polymers such aspolylactide-polyglycolide. Depending on the ratio of drug to polymer,and the nature of the particular polymer employed, the rate of drugrelease can be controlled. Examples of other biodegradable polymersinclude poly(orthoesters) and poly(anhydrides). Depot injectableformulations are also prepared by entrapping the drug in liposomes ormicroemulsions, which are compatible with body tissue.

III. Treatment Methods

In some embodiments, the present invention provides a compound ofstructure (I) or (II) for use as a medicament. For example, in oneaspect the present invention relates to a method of treating a diseaseassociated with cellular necrosis. In particular, one embodiment of theinvention provides methods for preventing or treating a disorderassociated with cellular necrosis (i.e., a necrotic cell disease) in amammal, comprising the step of administering to said mammal atherapeutically effective amount of a compound (e.g., compound ofstructure (I) or (II)) or therapeutic preparation of the presentinvention. Some other embodiments include use of a compound of structure(I) or (II) for treatment of a necrotic cell disease. Other embodimentsinclude use of a compound of structure (I) or (II) for manufacture of amedicament for treating a necrotic cell disease.

In certain embodiments, the disorder associated with cellular necrosisis a disorder such as trauma, ischemia, stroke, cardiac infarction,infection and/or sepsis. In certain embodiments, the disorder associatedwith cellular necrosis is a disorder such as trauma, ischemia, stroke,cardiac infarction, infection, Gaucher's disease, Krabbe disease and/orsepsis. In other embodiments, the disorder is a neurodegenerativedisease, such as Parkinson's disease (PD), Alzheimer's disease (AD),amyotrophic lateral sclerosis (ALS), Huntington's disease (HD), andHIV-associated dementia (HAD). In other embodiments the disorder is anischemic disease of organs including but not limited to brain, heart,kidney, and liver. In some different embodiments, the disorder is anocular disorder such as retinal degenerative disease, glaucoma orage-related macular degeneration. In some different embodiments, thedisorder is an autoimmune disorder such as rheumatoid arthritis,psoriasis or psoriatic arthritis. In some different embodiments, thedisorder is a central nervous system (CNS) disorder.

In other embodiments the presently disclosed compounds (e.g., compoundsof structure (I) or (II)) are used in methods for treatment ofinflammatory disorders. Some other embodiments are directed to use of acompound of structure (I) or (II) for treatment of an inflammatorydisorder. Other different embodiments include use of a compound ofstructure (I) or (II) for manufacture of a medicament for treating aninflammatory disorder. In some embodiments, the disorder is aninflammatory disease of the intestines such as Crohn's disease orulcerative colitis (both generally known together as inflammatory boweldisease).

In certain embodiments, the mammal is a primate, canine or felinesubject. In other embodiments, the mammal is a human subject. While notwishing to be bound by theory, it is believed that inhibition of RIP1kinase by the presently disclosed compounds is responsible, at least inpart, for their anti-inflammatory activity. Accordingly, embodiments ofthe invention also include methods for inhibiting RIP1 kinase, either invitro or in a subject in need thereof, the method comprises contacting aRIP1 kinase with a compound disclosed herein (e.g. compounds ofstructure (I) or (II)). In some of these embodiments, inhibiting RIP1kinase is effective to block (partially or fully) the release ofinflammatory mediators such as TNF and/or IL6.

The compounds of structure (I) and/or (II) can be used to treat ocularindications, for example to reduce or prevent the loss of photoreceptorand/or retinal pigment epithelial cell viability. In one aspect, theinvention provides a method of preserving the visual function of an eyeof a subject with an ocular condition, wherein a symptom of the ocularcondition is the loss of photoreceptor cell viability in the retina ofthe eye with the condition. The method comprises administering to theeye of the subject an effective amount of a compound of structure (I)and/or (II) thereby preserving the viability of the photoreceptor cellsdisposed within the retina of the eye. After administration of thecompound of structure (I) and/or (II) the visual function (e.g., visualacuity) of the eye may be preserved or improved relative to the visualfunction of the eye prior to administration of the compound of structure(I) and/or (II).

The ocular condition may be a condition selected from the groupconsisting of age-related macular degeneration (AMD), retinosispigmentosa (RP), macular edema, diabetic retinopathy, central areolarchoroidal dystrophy, BEST disease, adult vitelliform disease, patterndystrophy, myopic degeneration, central serous retinopathy, Stargardt'sdisease, Cone-Rod dystrophy, North Carolina dystrophy, infectiousretinitis, inflammatory retinitis, uveitis, toxic retinitis andlight-induced toxicity. AMD may be the neovascular or the dry form ofAMD. Retinal detachment may be a rhegmatogenous, a serous, or atractional retinal detachment.

In another aspect, the invention provides a method of preserving theviability of retinal pigment epithelial (RPE) cells within the retina ofa subject with an ocular condition, wherein a symptom of the ocularcondition is the loss of retinal pigment epithelial cells in the retinaof the eye with the condition. The method comprises administering to theeye of the subject an effective amount of a compound of structure (I)and/or (II) thereby preserving the viability of the retinal pigmentepithelial cells. The ocular condition may be selected from the groupconsisting of AMD, BEST disease, myopic degeneration, Stargardt'sdisease, uveitis, adult foveomacular dystrophy, fundus falvimaculatus,multiple evanescent white dot syndrome, serpiginous choroidopathy, acutemultifocal posterior placoid epitheliopathy (AMPPE), and other uveitisdisorders.

In another aspect, the invention provides a method of preserving theviability of photoreceptor cells disposed within a retina of a subjectwith an ocular condition selected from the group consisting of AMD, RP,macular edema, diabetic retinopathy, central areolar choroidaldystrophy, BEST disease, adult vitelliform disease, pattern dystrophy,myopic degeneration, central serous retinopathy, Stargardt's disease,Cone-Rod dystrophy, North Carolina dystrophy, infectious retinitis,inflammatory retinitis, uveitis, toxic retinitis and light-inducedtoxicity. The method comprises administering to the eye an effectiveamount of a compound of structure (I) and/or (II) thereby to preservethe viability of the photoreceptor cells disposed within the retina ofthe subject with a condition.

In another aspect, the invention provides a method of preserving theviability of photoreceptor cells disposed within a retina of a mammalianeye following retinal detachment. The method comprises administering acompound of structure (I) and/or (II) to the eye in which a region ofthe retina has been detached in amounts sufficient to preserve theviability of photoreceptor cells disposed within the region of thedetached retina.

In certain embodiments, the retinal detachment may be a rhegmatogenousretinal detachment, tractional retinal detachment, or serous retinaldetachment. In other embodiments, the retinal detachment may occur as aresult of a retinal tear, retinoblastoma, melanoma or other cancers,diabetic retinopathy, uveitis, choroidal neovascularization, retinalischemia, pathologic myopia, or trauma.

In another aspect, the invention provides a method of preserving visualfunction of an eye of a subject with an ocular condition selected fromthe group consisting of AMD, RP, macular edema, central areolarchoroidal dystrophy, retinal detachment, diabetic retinopathy, BESTdisease, adult vitelliform disease, pattern dystrophy, myopicdegeneration, central serous retinopathy, Stargardt's disease, Cone-Roddystrophy, North Carolina dystrophy, infectious retinitis, inflammatoryretinitis, uveitis, toxic retinitis and light-induced toxicity, whereina symptom of the ocular condition is the loss of photoreceptor cellsviability in the retina of the eye. The method comprises administeringan effective amount of a compound of structure (I) and/or (II) to thepatient.

In another aspect, the invention provides a method of preserving thevisual function of an eye of a subject with an ocular condition, whereina symptom of the ocular condition is the loss of photoreceptor cellviability and/or RPE viability in the retina of the eye. The methodcomprises treating the subject with a compound of structure (I) and/or(II); and (b), after treatment, measuring visual function (e.g., visualacuity) of the eye.

In other embodiments is provided a method of preserving the visualfunction of an eye of a subject with ocular conditions, wherein asymptom of the ocular condition is the loss of retinal ganglion cellviability in the retina of the eye with the conditions. The methodcomprises administering to the eye of the subject an effective amount ofa compound of structure (I) and/or (II) thereby preserving the viabilityof the retinal ganglion cells disposed within the retina of the eye.After administration of the compound of structure (I) and/or (II) thevisual function of the eye may be preserved or improved relative to thevisual function of the eye prior to administration of the compound ofstructure (I) and/or (II). Further, after the administration of thecompound of structure (I) and/or (II), the preserved retinal ganglioncell is capable of supporting axonal regeneration.

In each of the foregoing methods, the ocular condition, wherein asymptom of the condition is the loss of retinal ganglion cell viabilityin the retina of the eye, includes but is not limited to glaucoma, opticnerve injury, optic neuritis, optic neuropathies, diabetic retinopathy,central retinal artery occlusion, and central retinal vein occlusion. Itis contemplated that the forgoing methods may be used for the treatmentof optic neuropathies such as ischemic optic neuropathy (e.g., arteriticor non-arteritic anterior ischemic neuropathy and posterior ischemicoptic neuropathy), compressive optic neuropathy, infiltrative opticneuropathy, traumatic optic neuropathy, mitochondrial optic neuropathy(e.g., Leber's optic neuropathy), nutritional optic neuropathy, toxicoptic neuropathy, and hereditary optic neuropathy (e.g., Leber's opticneuropathy, Dominant Optic Atrophy, Behr's syndrome).

Also disclosed is a method of preserving the visual function of an eyeof a subject with an ocular condition selected from the group consistingof glaucoma, optic nerve injury, optic neuropathies, diabeticretinopathy, central retinal artery occlusion, and central retinal veinocclusion. The method comprises administering to the eye of the subjectan effective amount of a compound of structure (I) and/or (II) therebypreserving the viability of the retinal ganglion cells disposed withinthe retina of the eye and the visual function of the eye.

In another aspect, disclosed herein is a method of preserving theviability of retinal ganglion cells disposed within a retina of amammalian eye affected by, for example, glaucoma, optic nerve injury,optic neuritis, optic neuropathies, diabetic retinopathy, centralretinal artery occlusion, and central retinal vein occlusion. The methodcomprises administering a compound of structure (I) and/or (II) to theeye in which a region of the retina has been affected in amountssufficient to preserve the viability of retinal ganglion cells disposedwithin the region of the affected retina. The preserved retinal ganglioncell is capable of supporting axonal regeneration.

Also disclosed is a method for promoting axon regeneration in an eye ofa subject with an ocular condition, wherein a symptom of the ocularcondition is the loss of retinal ganglion cell viability in the retinaof the eye with the condition. The method comprises administering to theeye of the subject an effective amount of a compound of structure (I)and/or (II) thereby promoting axon regeneration of the retinal ganglioncell within the retina of the eye.

In each of the foregoing embodiments, it is understood that the methodsand compositions described herein can be used to preserve the viabilityand/or promote axon regeneration of retinal ganglion cells duringtreatment of the underlying conditions including, but not limited to,glaucoma, optic nerve injury, optic neuritis, optic neuropathies,diabetic retinopathy, central retinal artery occlusion, and centralretinal vein occlusion.

In another embodiment, the compound of structure (I) and/or (II) can beused to preserve neuron viability and promote axon growth and nervefunctions. Accordingly, the compound of structure (I) and/or (II) may beused to reduce or even reverse the loss of cognitive, motor, and sensoryfunctions associated with a CNS disorder, by preserving neuron viabilityand/or promoting axon regeneration and/or nerve functions.

In one aspect, the invention provides a method for promoting axonregeneration in a CNS neuron by exposing the CNS neuron to an effectiveamount of a compound of structure (I) and/or (II). The CNS neuron may beex vivo or in vivo. The CNS neuron may include, but is not limited to, aCNS sensory neuron, a motor neuron, a cortical neuron, a cerebellarneuron, a hippocampal neuron, and a midbrain neuron.

In another aspect, the invention provides a method for promoting nervefunction following injury to a CNS neuron. The method comprisesadministering to a subject an effective amount of a compound ofstructure (I) and/or (II) thereby to promote CNS neuron function. In afurther aspect, the invention provides a method for preserving theviability of a CNS neuron, wherein the method comprises administering toa subject an effective amount of a compound of structure (I) and/or (II)thereby to preserve the viability of the CNS neuron. Afteradministration of the compound of structure (I) and/or (II), the CNSneuron may be capable of supporting axonal regeneration.

In another aspect, the invention provides a method of treating a CNSdisorder in a subject in need thereof, wherein a symptom of the CNSdisorder is axon degeneration or injury within a CNS neuron. The methodcomprises administering to the subject an effective amount of a compoundof structure (I) and/or (II) thereby to promote regeneration of an axonin a CNS neuron affected by the CNS disorder. Following administrationof the compound of structure (I) and/or (II), neural functions may bemeasured, for example, as an indication of axon regeneration. It is alsocontemplated that, following administration of the compound of structure(I) and/or (II), the neuron function of the CNS neuron is preserved orimproved relative to the neuron function prior to administration of thecompound of structure (I) and/or (II). The CNS disorder includes, but isnot limited to, brain injury, spinal cord injury, dementia, stroke,Alzheimer's disease, amyotrophic lateral sclerosis (ALS/Lou Gehrig'sDisease), Parkinson's disease, Huntington's disease, multiple sclerosis,diabetic neuropathy, polyglutamine (polyQ) diseases, stroke, Fahrdisease, Menke's disease, Wilson's disease, cerebral ischemia, and aprion disorder. In exemplary embodiments, the CNS disorder is braininjury or spinal cord injury.

Also provided herein are methods for promoting neuron survival and axonregeneration in the CNS. CNS disorders characterized by impaired orfailing axon growth or axon degeneration may arise from CNS neuroninjury (e.g., trauma, surgery, nerve compression, nerve contusion, nervetransection, neurotoxicity, or other physical injury to the brain orspinal cord) or neurodegenerative CNS disease, wherein a symptom of thedisorder is axon degeneration (e.g., Alzheimer's disease, amyotrophiclateral sclerosis (ALS/Lou Gehrig's Disease), Parkinson's disease,multiple sclerosis, diabetic neuropathy, polyglutamine (polyQ) diseases,and stroke, Fahr disease, Menke's disease, Wilson's disease, cerebralischemia, prion disorder (e.g., Creutzfeldt-Jakob disease). In anexemplary embodiment, the CNS disorder is brain injury (e.g., traumaticbrain injury) or spinal cord injury (e.g., chronic, acute, or traumaticspinal cord injury). In another embodiment, the CNS disorder affects asubject's basic vital life functions such as breathing, heart beat andblood pressure, e.g., an injury to or aneurysm in the brain stem.

In certain embodiments, the CNS disorder affects a subject's cognitiveability, such as, brain injury to the cerebral cortex or aneurodegenerative CNS disorder, such as, Alzheimer's disease,frontotemporal dementia, dementia with Lewy bodies, corticobasaldegeneration, progressive supranuclear palsy, and prion disorders.

In other embodiments, the CNS disorder affects a subject's movementand/or strength, such as injury to the brain or spinal cord, or aneurodegenerative CNS disorder such as Parkinson's disease,frontotemporal dementia, dementia with Lewy bodies, corticobasaldegeneration, progress supranuclear palsy, Huntington's disease,multiple system atrophy, amyotrophic lateral sclerosis, and hereditaryspastic paresis.

In yet another embodiment, the CNS disorder affects a subject'scoordination, such as brain injury to the cerebellum or aneurodegenerative CNS disorder such as spinocerebellar atrophies,Friedreich's ataxia, and prion disorders.

In each of the foregoing methods, the CNS disorder includes, but is notlimited to, brain injury, spinal cord injury, Alzheimer's disease,amyotrophic lateral sclerosis (ALS/Lou Gehrig's Disease), Parkinson'sdisease, multiple sclerosis, diabetic neuropathy, polyglutamine (polyQ)diseases, stroke, Fahr disease, Menke's disease, Wilson's disease,cerebral ischemia, a prion disorder (e.g., Creutzfeldt-Jakob disease),dementia (e.g., frontotemporal dementia, dementia with Lewy bodies),corticobasal degeneration, progressive supranuclear palsy, multiplesystem atrophy, hereditary spastic paraparesis, and spinocerebellaratrophies.

It is understood that embodiments of the present invention include useof a compound of structure (I) or (II) for treatment of any of theforegoing indications. Other embodiments include use of a compound ofstructure (I) or (II) for manufacture of medicament for treatment of anyof the foregoing indications.

The term “trauma” as used herein refers to any physical damage to thebody caused by violence, accident, fracture etc. The term “ischemia”refers to a cardiovascular disorder characterized by a low oxygen stateusually due to the obstruction of the arterial blood supply orinadequate blood flow leading to hypoxia in the tissue. The term“stroke” refers to cardiovascular disorders caused by a blood clot orbleeding in the brain, most commonly caused by an interruption in theflow of blood in the brain as from clot blocking a blood vessel, and incertain embodiments of the invention the term stroke refers to ischemicstroke or hemorrhagic stroke. The term “myocardial infarction” refers toa cardiovascular disorder characterized by localized necrosis resultingfrom obstruction of the blood supply.

The phrase “therapeutically-effective amount” as used herein means thatamount of a compound, material, or composition comprising a compound ofthe present invention which is effective for producing some desiredtherapeutic effect in at least a sub-population of cells in an animal ata reasonable benefit/risk ratio applicable to any medical treatment. Atherapeutically effective amount for treating a neurological disorder isan amount sufficient to inhibit necrosis in at least a subset of cellsthat were exposed to a cell-death initiating event. Accordingly, atherapeutically effective amount prevents or minimizes diseaseprogression associated with cellular necrosis. Disease progression canbe monitored relative to an expected disease progression that is basedon population studies, controlled observations in individuals, or acombination of both.

In certain embodiments, a compound or pharmaceutical preparation isadministered orally. In other embodiments, the compound orpharmaceutical preparation is administered intravenously. Alternativeroutes of administration include sublingual, intramuscular, andtransdermal administrations.

In certain embodiments, the present invention relates to compounds forinhibiting cell death, wherein the compounds are represented bystructures (I) or (II). In certain embodiments, the compounds of thepresent invention are inhibitors of cell death. In any event, thecompounds of the present invention preferably exert their effect oninhibiting cell death at a concentration less than about 50 micromolar,more preferably at a concentration less than about 10 micromolar, andmost preferably at a concentration less than 1 micromolar.

The compounds of the invention can be tested in standard animal modelsof stroke and standard protocols such as described by Hara, H., et al.Proc Natl Acad Sci USA, 1997. 94(5): 2007-12.

When the compounds of the present invention are administered aspharmaceuticals, to humans and animals, they can be given per se or as apharmaceutical composition containing, for example, 0.1% to 99.5% (morepreferably, 0.5% to 90%) of active ingredient in combination with apharmaceutically acceptable carrier.

The preparations of the present invention may be given orally,parenterally, topically, or rectally. They are of course given in formssuitable for each administration route. For example, they areadministered in tablets or capsule form, by injection, inhalation, eyelotion, ointment, suppository, etc. administration by injection,infusion or inhalation; topical by lotion or ointment; and rectal bysuppositories. Oral administrations are preferred.

The phrases “parenteral administration” and “administered parenterally”as used herein means modes of administration other than enteral andtopical administration, usually by injection, and includes, withoutlimitation, intravenous, intramuscular, intraarterial, intrathecal,intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal,transtracheal, subcutaneous, subcuticular, intraarticulare, subcapsular,subarachnoid, intraspinal and intrasternal injection and infusion.

The phrases “systemic administration,” “administered systemically,”“peripheral administration” and “administered peripherally” as usedherein mean the administration of a compound, drug or other materialother than directly into the central nervous system, such that it entersthe patients system and, thus, is subject to metabolism and other likeprocesses, for example, subcutaneous administration.

These compounds may be administered to humans and other animals fortherapy by any suitable route of administration, including orally,nasally, as by, for example, a spray, rectally, intravaginally,parenterally, intracistemally and topically, as by powders, ointments ordrops, including buccally and sublingually.

Regardless of the route of administration selected, the compounds of thepresent invention, which may be used in a suitable hydrated form, and/orthe pharmaceutical compositions of the present invention, are formulatedinto pharmaceutically-acceptable dosage forms by conventional methodsknown to those of skill in the art.

Actual dosage levels of the active ingredients in the pharmaceuticalcompositions of this invention may be varied so as to obtain an amountof the active ingredient that is effective to achieve the desiredtherapeutic response for a particular patient, composition, and mode ofadministration, without being toxic to the patient.

The selected dosage level will depend upon a variety of factorsincluding the activity of the particular compound of the presentinvention employed, or the ester, salt or amide thereof, the route ofadministration, the time of administration, the rate of excretion ormetabolism of the particular compound being employed, the duration ofthe treatment, other drugs, compounds and/or materials used incombination with the particular compound employed, the age, sex, weight,condition, general health and prior medical history of the patient beingtreated, and like factors well known in the medical arts. A daily,weekly, or monthly dosage (or other time interval) can be used.

A physician or veterinarian having ordinary skill in the art can readilydetermine and prescribe the effective amount of the pharmaceuticalcomposition required. For example, the physician or veterinarian couldstart doses of the compounds of the invention employed in thepharmaceutical composition at levels lower than that required to achievethe desired therapeutic effect and then gradually increasing the dosageuntil the desired effect is achieved.

In general, a suitable daily dose of a compound of the invention will bethat amount of the compound that is the lowest dose effective to producea therapeutic effect (e.g., inhibit necrosis). Such an effective dosewill generally depend upon the factors described above. Generally dosesof the compounds of this invention for a patient, when used for theindicated effects, will range from about 0.0001 to about 100 mg per kgof body weight per day. Preferably the daily dosage will range from0.001 to 50 mg of compound per kg of body weight, and even morepreferably from 0.01 to 10 mg of compound per kg of body weight.

If desired, the effective daily dose of the active compound may beadministered as two, three, four, five, six or more sub-dosesadministered separately at appropriate intervals throughout the day,optionally, in unit dosage forms.

In another aspect of the invention the compounds can be administered incombination with other agents, including (but not limited to) compoundsthat are apoptosis inhibitors; PARP poly(ADP-ribose) polymeraseinhibitors; Src inhibitors; agents for the treatment of cardiovasculardisorders; anti-inflammatory agents, anti-thrombotic agents;fibrinolytic agents; anti-platelet agents, lipid reducing agents, directthrombin inhibitors; glycoprotein IIb/IIIa receptor inhibitors; calciumchannel blockers; beta-adrenergic receptor blocking agents;cyclooxygenase (e.g., COX-1 and COX-2) inhibitors; angiotensin systeminhibitor (e.g., angiotensin-converting enzyme (ACE) inhibitors); renininhibitors; and/or agents that bind to cellular adhesion molecules andinhibit the ability of white blood cells to attach to such molecules(e.g., polypeptides, polyclonal and monoclonal antibodies). Suitablecombination agents in this regard are also disclosed in U.S. Pat. No.7,491,743, and which is hereby incorporated by reference in its entiretyfor all that it discloses.

Embodiments of the invention also provide combinations of two or morecompounds that inhibit cellular necrosis (e.g., a compound as disclosedherein and an additional agent for inhibiting necrosis). The inventionalso provides combinations of one or more compounds that inhibitcellular necrosis combined with one or more additional agents orcompounds (e.g., other therapeutic compounds for treating a disease,condition, or infection such as an apoptosis inhibitor).

The invention also provides kits including one or more compounds orcombinations of the invention. A kit can also include one or moreadditional agents or compounds described herein. The differentcomponents of the kit can be provided in different containers. The kitcan be compartmentalized to receive the containers in close confinement.The kit can also contain instructions for using the compounds accordingto the invention.

As used herein, a kit such as a compartmentalized kit includes any kitin which compounds or agents are contained in separate containers.Illustrative examples of such containers include, but are not limitedto, small glass containers, plastic containers or strips of plastic orpaper. Particularly preferred types of containers allow the skilledworker to efficiently transfer reagents from one compartment to anothercompartment such that the samples and reagents are notcross-contaminated and the agents or solutions of each container can beadded in a quantitative fashion from one compartment to another. Suchcontainers include, but are not limited to, a container that will accepta compound or combination of compounds and/or other agents of theinvention. One or more compounds or agents can be provided as a powder(e.g. lyophilized powder) or precipitate. Such compound(s) can beresuspended prior to administration in a solution that may be providedas part of the kit or separately available. A kit can contain compoundsor agents in other forms such as liquids, gels, solids, as describedherein. Different compounds and/or agents may be provided in differentforms in a single kit.

The examples and preparations provided below further illustrate andexemplify the compounds of the present invention and methods for testingsuch compounds. It is to be understood that the scope of the presentinvention is not limited in any way by the scope of the followingexamples. In the following examples, and throughout the specificationand claims, molecules with a chiral center, unless otherwise noted,exist as a racemic mixture. Single enantiomers may be obtained bymethods known to those skilled in the art.

IV. Synthetic Methods

In other embodiments, the present disclosure provides improved methodsfor preparation of indole-hydantoin compounds, which in some embodimentsare potent inhibitors of necrosis. Although other syntheses of relatedcompounds have been proposed, none of these earlier methods issufficiently robust for large-scale manufacturing or for preparation andscreening of numerous diverse analogs. Specifically, U.S. Pat. No.7,491,743, which is hereby incorporated by reference in its entirety,provides three synthetic routes for these compounds. In the first ofthese routes, which is illustrated in Comparative Reaction Scheme 1a, anindole-3-carboxaldehyde fragment is connected to a hydantoin fragmentsby aldol reaction (Scheme 1a, step c). The target product is thenobtained by hydrogenation/reduction (Scheme 1a, step d). Among thedisadvantages of this method are that the obtained target product isachiral, and the substituted indole must be prepared via variousdifferent routes depending on the desired substitution.

A second approach to preparation of the indole-hydantoin compoundsdescribed in U.S. Pat. No. 7,491,743 is illustrated in ComparativeReaction Scheme 2. In this scheme, the aldehyde was obtain by reduction,iodination, cyano-substitution and cyano-reduction. The target productwas then obtained by cyclization in the presence of ammonium carbonateand potassium cyanide, followed by alkylation. Because of the achiralproducts, linear synthetic strategy and toxicities of the potassiumcyanide, this route is also not conductive to divergent synthesismethods and large-scale production.

Comparative Reaction Scheme 3 illustrates the third method described inU.S. Pat. No. 7,491,743. In this approach, the N-acetyl tryptophanderivatives were obtained by mixing indoles with serine in acetic acidand acetic anhydride. The target product was then obtained by kineticresolution, methylation, amination and finally reaction withtriphosgene. Although this method does provide chiral products, it isinefficient due to the long synthetic route and low reaction yields (thereaction with serine only yield 40% product and cyclization bytriphosgene yield 34%), and also requires use of enzymes to obtain thedesired chiral purity.

In contrast to the above methods, the presently described methods areamendable to high-yielding, large scale production without requiringkinetic resolution. Accordingly, in some embodiments the inventionprovides a method for preparing a compound having the followingstructure (III):

or a pharmaceutically acceptable salt, prodrug, stereoisomer or tautomerthereof, wherein:

R¹, R², R³ and R⁴ are each independently H, halo or C₁-C₆ alkyl;

R⁵ is H or halo; and

R⁶ is H, C₁-C₆ alkyl, C₁-C₆ alkenyl, aralkyl, cycloalkylalkyl orheterocyclylalkyl, the method comprising reacting aldehyde (5), or asalt, stereoisomer or tautomer thereof, with phenylhydrazine (6), or asalt thereof, to yield (7), or a salt, stereoisomer or tautomer thereof,according to Reaction Scheme 1:

wherein each P is independently H or a protecting group. In someembodiments, each P is H. In other embodiments, each P is a protectinggroup such as butyloxycarbonyl (Boc).

In exemplary embodiments of Reaction Scheme 1, compound 5 is solvated inan alcohol solvent (e.g., methanol or ethanol) and compound 6 is thenbrought into contact with solvated compound 5. After contacting (e.g.,stirring) for an amount of time sufficient for the reaction, an acid,such as sulfuric or phosphoric acid is added to the mixture to yieldcompound 7. In various embodiments, the mixture of 5 and 6 is stirred attemperatures of about 10-50° C., or about 10-30° C., before addition ofthe acid. In some other embodiments, the reaction temperature isincreased to about 95-100° C. for a sufficient period of time afteraddition of the acid.

Compounds 5 and 6 can be present in various concentrations within themixture of compounds 5 and 6 according to Reaction Scheme 1. Forexample, in some embodiments, the molar ratio of compound 5 to 6 rangesfrom about 1:0.9 to about 1:1.1, for example about 1:1. Compound 5 istypically present in about 1 gram for every 10-20 mL of solvent.

Different enantiomers of compound (5) can be used depending on thedesired stereochemistry of the final product. Accordingly, in someembodiments aldehyde (5) has one of the following structures (5′) or(5″):

As will be apparent to one of skill in the art, the stereochemistry ofcompound (7) will result from the stereochemistry of compound (5) andother precursors. In some embodiments, compound (7) has one of thefollowing structures (7′) or (7″):

The disclosed methods are useful for preparation of compounds ofstructure (III) having various different R⁶ groups. For example, in someembodiments R⁶ is C₁-C₆ alkyl, C₁-C₆ alkenyl, aralkyl, cycloalkylalkylor heterocyclylalkyl, and the method further comprises reacting compound(7), or a salt, stereoisomer or tautomer thereof, with an alkylatingagent (8) to yield (1), or a salt, stereoisomer or tautomer thereof,according to Reaction Scheme 2:

wherein L is a leaving group.

Again, the stereochemistry of the final product is typically controlledby the stereochemistry of the precursors, and in various embodimentscompound (1) has one of the following structures (1′) or (1″):

An alkylating agent known in the art can be employed in the methods. Invarious embodiments the leaving group is halogen, for example bromine oriodine. In some different embodiments, the leaving group is sulfonate,such as p-toluenesulfonate or triflate.

Conditions for alkylation of compound 7 according to Reaction Scheme 2are readily determined by one of ordinary skill in the art. In someembodiments, compound 7 is dissolved in a solvent, the alkylationreagent (L-R⁶) and a base are added to the mixture, then the reactionmixture is stirred. Exemplary solvents for this purpose include polarand non-protic solvents, for example DMSO or DMF. The amount of thesolvent can be varied, and in certain embodiments the solvent is presentin about 10-40 mL/g based on compound 7.

Some embodiments include use of a base to effect the reactionillustrated in Reaction Scheme 2. In some embodiments, the base is analkali metallic carbonate or tert-alkylamine having from 1-4 carbonatoms. In certain embodiments, the alkali metallic carbonate ispotassium carbonate, cesium carbonate or sodium carbonate. In otherembodiments, the tert-alkylamine is triethylamine. The amount of baseemployed can be varied, and typically ranges from about 1.2-1.5 molesper mole of compound 7, for example about 1.5 mole per mole of compound7.

In various embodiments, the alkylation reagent is present at about1.2-1.5 moles per mole of compound 7, for example about 1.5 moles permole of compound 7. In other various embodiments, the alkylation ofcompound (7) is performed at temperatures from about 0-80° C., forexample about 10-30° C. In other embodiments, the alkylation reagent(L-R⁶) is methyl p-toluensulfonate or L-methyl, wherein L is a halogen.

Compounds with different R⁶ moieties can be prepared by selection of theappropriate alkylating agent. In some embodiments, R⁶ is C₁-C₆ alkyl,for example in some embodiments R⁶ is methyl. In other embodiments, R⁶is ethyl, propyl, isopropyl or butyl. Embodiments wherein R⁶ is methyltypically employ methyl iodide as the alkylating reagent. In someembodiments, the C₁-C₆ alkyl is substituted. In other embodiments, theC₁-C₆ alkyl is unsubstituted.

In various different embodiments, R⁶ is aralkyl. In some embodiments,the aralkyl is substituted. In other embodiments, the aralkyl isunsubstituted. The aryl moiety of the aralkyl can be any of a number ofaryl moieties, including phenyl, biphenyl and phenol moieties.

In some specific embodiments, R⁶ is benzyl. For example, in someembodiments R⁶ has the following structure:

wherein R^(6a), R^(6b), R^(6c) and R^(6d) are each independently H,halo, C₁-C₆ alkyl or C₁-C₆ haloalkyl. In some embodiments, R^(6a) is H,F or Cl; R^(6b) is H, F, Cl, CH₃ or CF₃; R^(6c) is H, F, Cl, I or CH₃;and R^(6d) is H, F or CF₃.

In some other specific embodiments, R⁶ has one of the followingstructures:

In other different embodiments, R⁶ is heterocyclyalkyl. In some of theseembodiments, the hetereocyclylalkyl is substituted. In some other ofthese embodiments, the hetereocyclylalkyl is unsubstituted. In someexemplary embodiments, the heterocyclylalkyl is heteroarylalkyl. Forexample, in various embodiments R⁶ is pyridinylalkyl, for example thefollowing structure:

In other embodiments, the heterocyclylalkyl is aliphatic, for example insome embodiments the heterocycle of the heterocyclylalkyl comprisesoxygen or nitrogen, for example tetrahydrofuranyl, pyranyl, piperidinylor ethylene oxide. In some embodiments, the heterocyclylalkyl has one ofthe following structures:

In some different embodiments, R⁶ is cycloyalkylalkyl, such ascyclopropyl or cyclopentyl. For example, one of the followingstructures:

In some different embodiments, R⁶ is C₁-C₆ alkenyl. For example, in someembodiments R⁶ has the following structure:

Compounds of structure (III), wherein R⁵ is halogen (i.e., R⁵) can beprepared in various embodiments by methods which further comprisetreating compound (1) with a halogenating reagent to form a compound ofstructure (III), wherein R⁵ is halogen. Exemplary halogenating reagentin some embodiments are N-bromosuccinimide or N-chlorosuccinimide, whichcan be employed at molar ratios derivable by one of ordinary skill inthe art.

In some embodiments, the method for preparing compounds of structure(III), wherein R5 is halogen, comprise dissolving compound (III)(prepared according to Reaction Scheme 2) in a solvent under inertatmosphere (e.g., N₂), adding the halogenation reagent heating themixture to reflux for a sufficient period of time to effect halogenation(e.g., from about 3-6 hours).

Typical solvents for the halogenation of compound (III) includenon-polar solvents, such as CCl₄. The amount of the solvent used can bevaried according to the knowledge of one of ordinary skill in the art.

The halogenated product can be purified according to common procedures.In some embodiments, the reaction mixture is cooled to room temperatureand filtered. The filtrate is then concentrated and purified by flashchromatography. The procedure and mobile phase of flash chromatographyis selected according to the general practice in the art.

In some embodiments of the foregoing method, aldehyde (5) is provided ina protected form. The protecting group may be removed prior to reactionwith compound (6) according to Reaction Scheme 1. Exemplary deprotectionconditions include reduction, such as hydrogenation. Accordingly, insome embodiments aldehyde compound (5) has been prepared by reducingcompound (4):

wherein R′ is C₁-C₆ alkyl. In some embodiments, R′ is ethyl (Et).

Again, stereochemistry of the final product may depend on thestereochemistry of the intermediate compounds, and thus in someembodiments compound (4) has one of the following structures (4′) or(4″):

In certain embodiments, compound (5) is prepared from compound (4) underconditions for hydrogenation, for example by treating compound (4) withpalladium and triethylsilane. Typically the Pd is Pd/C (e.g., 10% ofPd/C). In some embodiments, the ratio of the volume of triethylsilane tothe mass of Pd/C ranges from about 10:1 to about 30:1, for example about20:1. In certain embodiments, the ratio of triethylsilane to compound 4ranges from about 1:1 to about 5:1, for example about 3:1.

Various different conditions familiar to those of skill in the art canbe used for reduction of compound 4. For example, in some embodimentscompound (4) is dissolved in a solvent and cooled in an ice bath,followed by addition of the reducing agent. In some exemplaryembodiments, the reaction mixture thus obtained is stirred in the icebath for 3-5 min. and warmed to room temperature for 0.5-1 hours toprovide compound 5. Typical reaction temperatures for the reduction ofcompound (4) range from about −10-40° C., for example about 0-30° C.

Typical solvents for reduction of compound (4) include tetrahydrofuranand dichloromethane. The amount of the solvent can be varied and is someembodiments ranges from about 10-40 mL/g based on compound 4.

In still other embodiments, compound (4) has been prepared from thecorresponding acid (3) according to Reaction Scheme 3:

wherein AA is an acid activating reagent, such as DCC, EDCI, or otheractivating agents known in the art.

In some more specific embodiments, compound (3) has one of the followingstructures (3′) or (3″):

For the synthesis of compound 4, typical procedures comprise dissolvingcompound (3) in a solvent and cooling in an ice bath. The activating(condensation) reagent, alkylthiol (e.g., ethanethiol) and an optionalcatalyst are then added. The reaction mixture is then stirred in the icebath for 5-15 min (e.g., 10 min) and then warmed to room temperature for2-3 hours to provide compound 4.

In still further embodiments of the foregoing method compound (3) hasbeen prepared from an amino acid having the following structure (2):

In various embodiments, compound (2) has one of the following structures(2′) or (2″):

In still more embodiments, the method comprises one or more of thefollowing transformations (A), (B), (C), (D), (E) or (F):

wherein:

each P is independently H or a protecting group;

each P¹ is independently a protecting group; and

and L is a leaving group.

In some embodiments of the foregoing, the method comprisestransformation (E) and least one of transformations (A), (B), (C), (D)and/or (F). In various embodiments, the method comprises transformation(D) and (E). In other embodiments, the method comprises each oftransformations (A), (B), (C), (D), (E) and (F).

In still other embodiments, the method further comprises the followingtransformation:

wherein R⁵′ is halo.

The synthetic methods described herein may be used to prepare variousindole-hydantoin compounds of structure (III). In some embodiments, thecompounds are compounds wherein R¹ is H, Cl or F. In other embodiments,R² is H, Cl or F. In still more embodiments, R³ is H, Cl or F. In otherembodiments, R⁴ is H, Cl or I. In some more specific embodiments, R¹ isH, R² is H, R³ is F and R⁴ is Cl. In still other embodiments, Each ofR¹, R^(2′) and R³ are H and R⁴ is Cl. In various embodiments of any ofthe foregoing, R⁶ is methyl, and in other embodiments, R⁶ is benzyl,which is unsubstituted or substituted with one or more halo.

In still other embodiments R⁵ is H. In different embodiments, R⁵ is Bror Cl.

Exemplary compounds which can be prepared according to variousembodiments of the methods include the following compounds:

Compounds useful in the above synthetic methods are also provided.Accordingly, in some embodiments, the invention provides a compoundhaving one of the following structures (4′) or (4″):

or a salt or tautomer thereof, wherein:

each P is independently H or a protecting group; and

R′ is C₁-C₆ alkyl, such as ethyl.

Mixtures (e.g., racemic mixtures) of (4′) and (4″) or (5′) and (5″) arealso provided.

It will also be appreciated by those skilled in the art that in theprocess described herein the functional groups of intermediate compoundsmay need to be protected by suitable protecting groups. Such functionalgroups include hydroxy, amino, mercapto and carboxylic acid. Suitableprotecting groups for hydroxy include trialkylsilyl or diarylalkylsilyl(for example, t-butyldimethylsilyl, t-butyldiphenylsilyl ortrimethylsilyl), tetrahydropyranyl, benzyl, and the like. Suitableprotecting groups for amino, amidino and guanidino includet-butoxycarbonyl, benzyloxycarbonyl, and the like. Suitable protectinggroups for mercapto include —C(O)—R″ (where R″ is alkyl, aryl orarylalkyl), p-methoxybenzyl, trityl and the like. Suitable protectinggroups for carboxylic acid include alkyl, aryl or arylalkyl esters.Protecting groups may be added or removed in accordance with standardtechniques, which are known to one skilled in the art and as describedherein. The use of protecting groups is described in detail in Green, T.W. and P. G. M. Wutz, Protective Groups in Organic Synthesis (1999), 3rdEd., Wiley. As one of skill in the art would appreciate, the protectinggroup may also be a polymer resin such as a Wang resin, Rink resin or a2-chlorotrityl-chloride resin.

It will also be appreciated by those skilled in the art, although suchprotected derivatives of compounds of this invention may not possesspharmacological activity as such, they may be administered to a mammaland thereafter metabolized in the body to form compounds of theinvention which are pharmacologically active. Such derivatives maytherefore be described as “prodrugs”. All prodrugs of compounds of thisinvention are included within the scope of the invention.

Furthermore, all compounds of the invention which exist in free base oracid form can be converted to their pharmaceutically acceptable salts bytreatment with the appropriate inorganic or organic base or acid bymethods known to one skilled in the art. Salts of the compounds of theinvention can be converted to their free base or acid form by standardtechniques.

EXAMPLES

All non-aqueous reactions were carried out in oven- or flame-driedglassware under nitrogen atmosphere. All chemicals were purchased fromcommercial vendors and used as is, unless otherwise specified. Reactionswere magnetically stirred and monitored by thin layer chromatography(TLC) with 250 μm pre-coated silica gel plates, visualized either withUV, or in an iodine chamber. Flash column chromatography was performedusing silica gel (100-200 mesh). Chemical shifts are reported relativeto chloroform (δ7.26), methanol (δ3.31), or DMSO (δ2.50) for ¹H NMR.

Example 1 Preparation of(R)-5-((7-chloro-6-fluoro-1H-indol-3-yl)methyl)-3-methylimidazolidine-2,4-dione(Ia)

(R)-3-(2,5-dioxoimidazolidin-4-yl)propanoic acid

NaOH (12.5 M, 80 mL, 1.0 mol) was added dropwise to a suspension ofD-glutamic acid (147 g, 1.0 mol) in water (120 mL) at 10° C. The mixturewas stirred at room temperature for 10 minutes, and then NaOCN (71.6 g,1.1 mol) was added. The reaction was heated to 80° C. for 3 hours.Concentrated HCl (183 mL, 2.2 mol) was added while maintaining atemperature below 20° C. by ice bath. The resulting mixture was refluxedfor 3 hours, then cooled to room temperature without stirring for 12hours until a white solid was deposited. The mixture was filtered andwashed with water (100 mL×2) and dried in vacuo to afford the desiredproduct as a white solid (122 g, 71%, 99% ee).

S-ethyl (R)-3-(2,5-dioxoimidazolidin-4-yl)propanethioate

To a solution of (R)-3-(2,5-dioxoimidazolidin-4-yl)propanoic acid (0.1mol, 17.2 g) in 200 mL DCM, EDC (0.11 mole, 21 g), EtSH (0.1 mol, 6.2g), and DMAP (0.008 mol, 0.97 g) were added sequentially while coolingthe reaction mixture to 0° C. with an ice bath. The reaction temperaturewas maintained for 10 minutes, followed by removal of the ice bath toallow the reaction temperature to warm to room temperature for 3 hours.The reaction was monitored by TLC (SiO₂, 100% methanol). Uponcompletion, the reaction was quenched by adding 0.5 M HCl (100 mL). Thecrude reaction mixture was then filtered and the aqueous phase wasextracted with DCM (100 mL×2). The organic phase was concentrated invacuo to yield a white solid. The filter cake was slurried in water (200mL) to remove ethanethiol. The slurry was filtered and dried over thefilter cake to afford the desired product as a white solid (17.5 g,81%). ¹H NMR (500 MHz, DMSO-d₆) δ 10.63 (s, 1H), 7.94 (s, 1H), 4.09-3.91(m, 1H), 2.82 (q, J=7.4 Hz, 2H), 2.74-2.60 (m, 2H), 2.03-1.90 (m, 1H),1.82-1.70 (m, 1H), 1.16 (t, J=7.4 Hz, 3H).

(R)-3-(2,5-dioxoimidazolidin-4-yl)propanal

To a solution of S-ethyl(R)-3-(2,5-dioxoimidazolidin-4-yl)propanethioate (10 mmol) in THF (20mL) cooled with an ice bath to 0° C., 10% Pd/C (0.2 g) and Et₃SiH (4.1mL, 30 mmol) were added. The ice bath was removed after 5 minutes, andthe reaction was allowed to stir at room temperature for 1 h while beingmonitored by TLC (SiO₂, DCM/methanol=10:1). Solid Pd/C was removed byfiltration and the crude reaction mixture was concentrated in vacuo toafford the desired product. The crude product was used in the nextsynthetic step without further purification.

(R)-5-((7-chloro-6-fluoro-1H-indol-3-yl)methyl)imidazolidine-2,4-dione

To a solution of (R)-3-(2,5-dioxoimidazolidin-4-yl)propanal (10 mmol) inmethanol (20 mL) (2-chloro-3-fluorophenyl)hydrazine hydrochloride (10mmol) was added. The reaction was allowed to stir at room temperaturefor 2 hours while being monitored by TLC (SiO₂, DCM/methanol=10:1). Whenthe reaction was complete, methanol was removed in vacuo, affording thecrude product as a dark brown solid. The mixture was then refluxed for 2hours in 10% H₂SO₄ (20 mL) while monitoring the reaction by TLC (SiO₂,DCM/methanol=10:1). After the reaction was complete, the mixture wasallowed to cool to room temperature. The mixture was extracted withethyl acetate (4×10 mL) and the combined organic phases were pooled anddried over anhydrous Na₂SO₄. The product was filtered and concentratedin vacuo and purified by column chromatography (SiO₂) and recrystallizedto afford the desired product (0.85 g, 33% 99.3% ee). ¹H NMR (400 MHz,DMSO-d₆) δ 11.45 (s, 1H), 10.37 (s, 1H), 7.90 (s, 1H), 7.52 (dd, J=8.7,4.7 Hz, 1H), 7.21 (d, J=2.4 Hz, 1H), 7.03 (dd, J=10.2, 8.8 Hz, 1H), 4.33(t, J=4.5 Hz, 1H), 3.06 (dd, J=4.4, 3.1 Hz, 2H); ¹³C NMR (126 MHz,DMSO-d₆) δ 176.08, 157.82, 155.12, 153.23, 133.46, 126.06, 118.75,110.09, 108.18, 102.27, 58.58, 26.67; ESI-MS m/z 282.4 (M+H)⁺;MALDI-HRMS m/z calcd for C₁₂H₉ClFN₃O₂(M)⁺ 281.0367. found 281.0361.

(R)-5-((7-chloro-6-fluoro-1H-indol-3-yl)methyl)-3-methylimidazolidine-2,4-dione

To a solution of(R)-5-((7-chloro-6-fluoro-1H-indol-3-yl)methyl)imidazolidine-2,4-dione(1 mmol) in 2 mL DMSO, methyl iodide (1.5 mmol) and K₂PO₄ (1.5 mmol)were added. The reaction mixture was stirred at room temperature for 2.5hours while monitoring by TLC (SiO₂, DCM/methanol=10:1). When thereaction was complete, it was quenched with 5 mL of water and extractedwith ethyl acetate (3×5 mL). The organic phases were pooled and driedover anhydrous Na₂SO₄. The product was filtered, concentrated in vacuo,and purified using column chromatography (SiO₂) to afford the desiredproduct (75%, 98.9% ee). ¹H NMR (500 MHz, DMSO-d₆) δ 11.48 (s, 1H), 8.22(d, J=10.6 Hz, 1H), 7.50 (dd, J=8.7, 4.7 Hz, 1H), 7.20 (d, J=2.4 Hz,1H), 7.02 (dd, J=10.1, 8.7 Hz, 1H), 4.35 (t, J=5.6 Hz, 1H), 3.11 (dd,J=15.0, 4.6 Hz, 1H), 3.05 (dd, J=14.9, 5.6 Hz, 1H), 2.62 (s, 3H); ¹³CNMR (126 MHz, DMSO-d₆) δ 174.52, 157.33, 155.10, 153.22, 133.52, 126.45,118.66, 110.08, 108.33, 102.45, 57.44, 26.89, 24.27; ESI-MS m/z 318.1(M+Na)⁺; ESI-HRMS m/z calcd for C₁₃H₁₁ClFN₃NaO₂ (M+Na)⁺ 318.0416. found318.0428.

Example 2 Exemplary Preparations of Compound 4

To a solution of compound 3 (0.1 mol, 17.2 g) in 200 mL DCM, EDCI (0.11mol, 21 g) in the ice bath (0-5° C.), EtSH (0.1 mol, 6.2 g) and thecatalyst DMAP (0.008 mol, 0.97 g) were added sequentially. After keepingthe temperature for 10 min, the bath was removed and kept stirring atroom temperature for 2.5-3 h until TLC detected the reaction wascomplete (eluting with MeOH). The reaction was then quenched by adding100 mL 0.5 mol/L HCl solution. The reaction mixture was then filtratedafter stirring for 2 min and the filtrate was separated, and the aqueousphase was then extracted twice with 100 mL DCM. A white solid wasobtained from the combined organic phases and concentrated in vacuum.The solid and the filter cake was slurried in 200 mL water to removeethanethiol. The final product was obtained by filtering and drying overthe filter cake. (17.5 g, 81%), ¹H NMR (500 MHz, dmso-d₆) δ 10.63 (s,1H), 7.94 (s, 1H), 4.09-3.91 (m, 1H), 2.82 (q, J=7.4 Hz, 2H), 2.74-2.60(m, 2H), 2.03-1.90 (m, 1H), 1.82-1.70 (m, 1H), 1.16 (t, J=7.4 Hz, 3H).

Compound 4 was obtained according to the methods and conditionsdescribed above, with different condensation reagent and solvent. Theresults are summarized in Table 2 below:

TABLE 2 Exemplary Conditions for Preparation of Compound (4)Condensation Entry solvent reagent yield % 1 DCM DCC 65 2 DCM EDCl 81 3THF EDCl 72 4 THF DCC 70

Example 3 Preparation of Compound 5

To a solution of thioester compound 4 (10 mmol) in 20 mL THF in icebath, 10% Pd/C (0.2 g) and Et₃SiH (4.1 mL, 30 mmol) was added. The icebath was removed after 5 min and the reaction was allowed to stir atroom temperature for 0.5-1 h until TLC detected the reaction wascomplete (eluting with DCM:MeOH=10:1). The catalyst was removed byfiltration and the filtrate concentrated to provide compound 5.

Example 4 Exemplary Preparations of Compound 7

To a solution of compound 5 (10 mmol) in 20 mL MeOH in 100 mL roundflask, substituted phenylhydrazine hydrochloride 6 (1 equiv) was added.The reaction mixture was allowed to stir at room temperature for 1.5-2 huntil TLC detected the reaction was complete (eluting withDCM:MeOH=10:1). MeOH was removed to afford a dark brown solid and themixture was then refluxed for 2 h with 20 mL 10% H₂SO₄ until TLCdetected the reaction complete (eluting with DCM:MeOH=10:1). Aftercooling to room temperature, the reaction mixture was extracted withethylacetate (10 mL×4). The organic phases were combined and dried overanhydrous Na₂SO₄. After concentrating in vacuum, compound 7a wasobtained by column chromatography, which after recrystallization (0.85g, 33%) had an ee value: 99.3%. Analogous conditions produced 7b with anee value of 99.3%

Synthetic conditions and results for various compounds of structure (7)are provided in Table 3 below (the yield in this table is the yield fromcompound 4 to 7):

TABLE 3 Exemplary Preparations of Compound (7) entry 6 Product 7 yield %1

  6a

  7a 35 2

  6b

  7b 40 3

  6c

  7c 36 4

  6d

  7d 33 5

  6e

  7e 35 6

  6f

  7f 30 7

  6g

  7g 33 8

  6h

  7h 35 9

  6i

  7i 32Compounds 7a-i were analyzed by NMR and mass spectrometry. Data isprovided below:7a: ¹H NMR (500 MHz, DMSO-d₆) δ 11.27 (s, 1H), 10.37 (s, 1H), 7.90 (s,1H), 7.53 (d, J=7.9 Hz, 1H), 7.21 (d, J=2.3 Hz, 1H), 7.14 (d, J=7.5 Hz,1H), 6.98 (t, J=7.8 Hz, 1H), 4.33 (t, J=4.8 Hz, 1H), 3.10 (dd, J=14.0,4.0 Hz, 1H), 3.06 (dd, J=14.0, 3.6 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ176.10, 157.83, 133.11, 129.97, 126.01, 120.85, 119.85, 118.26, 116.15,109.88, 58.63, 26.90; ESI-MS m/z 286.1 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₂H₁₀ClN₃NaO₂(M+Na)⁺ 286.0354. found 283.0863.7b: ¹H NMR (500 MHz, DMSO-d₆) δ 10.85 (s, 1H), 10.36 (s, 1H), 7.89 (s,1H), 7.58 (d, J=7.9 Hz, 1H), 7.46 (d, J=7.4 Hz, 1H), 7.19 (d, J=2.5 Hz,1H), 6.80 (t, J=7.7 Hz, 1H), 4.33 (t, J=4.8 Hz, 1H), 3.07 (dd, J=15.1,5.1 Hz, 1H), 3.03 (dd, J=15.1, 4.8 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ176.12, 157.83, 138.02, 130.27, 128.56, 125.75, 120.84, 119.30, 109.99,77.27, 58.62, 26.99; ESI-MS m/z 356.2 (M+H)⁺; MALDI-HRMS m/z calcd forC₁₂H₁₀IN₃O₂354.9818 (⁷⁹I, M+H)⁺. found 354.9809.7c: ¹H NMR (500 MHz, DMSO-d₆) δ 10.74 (s, 1H), 10.37 (s, 1H), 7.87 (s,1H), 7.32 (s, 1H), 7.20 (d, J=8.2 Hz, 1H), 7.07 (d, J=2.3 Hz, 1H), 6.88(dd, J=8.2, 1.4 Hz, 1H), 4.29 (t, J=4.4 Hz, 1H), 3.05 (dd, J=14.0, 3.7Hz, 1H), 3.01 (dd, J=14.1, 4.5 Hz, 1H), 2.37 (s, 3H); ¹³C NMR (126 MHz,DMSO-d₆) δ 176.24, 157.87, 134.73, 128.17, 127.10, 124.60, 122.91,118.61, 111.41, 107.94, 58.81, 27.04, 21.83; ESI-MS m/z 244.2 (M+H)⁺;MALDI-HRMS m/z calcd for C₁₃H₁₄N₃O₂(M)⁺243.1016. found 244.1080.7d: ¹H NMR (500 MHz, DMSO-d₆) δ 11.48 (s, 1H), 10.36 (s, 1H), 7.90 (s,1H), 7.53 (d, J=8.5 Hz, 1H), 7.24 (d, J=1.4 Hz, 1H), 7.17 (d, J=8.5 Hz,1H), 4.33 (t, J=4.4 Hz, 1H), 3.09 (dd, J=15.4, 5.3 Hz, 1H), 3.05 (dd,J=15.5, 4.9 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 176.05, 157.80,134.01, 128.41, 126.94, 123.91, 120.62, 119.20, 114.34, 110.32, 58.58,26.67; ESI-MS m/z 296.0 (M−H)⁻; ESI-HRMS m/z calcd forC₁₂H₈Cl₂N₃O₂(M−H)⁻ 295.9999. found 295.9999.7e: ¹H NMR (500 MHz, DMSO-d₆) δ 11.52 (s, 1H), 10.39 (s, 1H), 7.92 (s,1H), 7.61 (d, J=1.7 Hz, 1H), 7.28 (d, J=2.5 Hz, 1H), 7.20 (d, J=1.8 Hz,1H), 4.34 (t, J=4.7 Hz, 1H), 3.11 (dd, J=14.9, 4.8 Hz, 1H), 3.05 (dd,J=15.0, 4.5 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ176.12, 157.81, 131.89,130.40, 127.86, 123.66, 120.47, 117.91, 116.89, 109.86, 58.65, 26.58;ESI-MS m/z 298.1 (M+H)⁺; MALDI-HRMS m/z calcd forC₁₂H₉Cl₂N₃O₂(M)⁺297.0072. found 297.0065.7f: ¹H NMR (500 MHz, DMSO-d₆) δ 11.65 (s, 1H), 10.61 (s, 1H), 7.89 (s,1H), 7.34 (d, J=2.1 Hz, 1H), 7.12 (d, J=8.1 Hz, 1H), 7.01 (d, J=8.1 Hz,1H), 4.33 (ddd, J=8.3, 4.4, 1.0 Hz, 1H), 3.49 (dd, J=14.8, 4.3 Hz, 1H),3.11 (dd, J=14.9, 8.4 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 175.89,157.93, 134.66, 127.86, 125.35, 124.11, 121.50, 120.66, 115.60, 111.02,59.34, 28.85; ESI-MS m/z 298.1 (M+H)⁺; MALDI-HRMS m/z calcd forC₁₂H₉Cl₂N₃O₂(M)⁺ 297.0072. found 297.0066.7g: ¹H NMR (400 MHz, DMSO-d₆) δ 11.45 (s, 1H), 10.37 (s, 1H), 7.90 (s,1H), 7.52 (dd, J=8.7, 4.7 Hz, 1H), 7.21 (d, J=2.4 Hz, 1H), 7.03 (dd,J=10.2, 8.8 Hz, 1H), 4.33 (t, J=4.5 Hz, 1H), 3.06 (dd, J=4.4, 3.1 Hz,2H); ¹³C NMR (126 MHz, DMSO-d₆) δ 176.08, 157.82, 155.12, 153.23,133.46, 126.06, 118.75, 110.09, 108.18, 102.27, 58.58, 26.67; ESI-MS m/z282.4 (M+H)⁺; MALDI-HRMS m/z calcd for C₁₂H₆ClFN₃O₂(M)⁺ 281.0367. found281.0361.7h: ¹H NMR (500 MHz, DMSO-d₆) δ 11.41 (s, 1H), 10.38 (s, 1H), 7.91 (s,1H), 7.35 (dd, J=9.7, 2.2 Hz, 1H), 7.27 (d, J=2.5 Hz, 1H), 7.11 (dd,J=9.2, 2.3 Hz, 1H), 4.33 (t, J=4.5 Hz, 1H), 3.09 (dd, J=14.9, 4.8 Hz,1H), 3.03 (dd, J=15.0, 4.5 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 176.16,157.80, 155.52, 130.17, 129.38, 128.12, 116.21, 110.19, 109.67, 103.59,58.65, 26.67; ESI-MS m/z 282.4 (M+H)⁺; MALDI-HRMS m/z calcd forC₁₂H₆ClFN₃O₂(M)⁺ 281.0367. found 281.0361.7i: ¹H NMR (500 MHz, DMSO-d₆) δ 11.57 (s, 1H), 10.53 (s, 1H), 7.89 (s,1H), 7.24 (d, J=2.3 Hz, 1H), 7.10 (dd, J=8.3, 4.1 Hz, 1H), 6.77 (dd,J=10.8, 8.3 Hz, 1H), 4.29 (ddd, J=7.2, 4.4, 1.0 Hz, 1H), 3.22 (dd,J=14.8, 4.3 Hz, 1H), 3.05 (dd, J=14.8, 7.4 Hz, 1H); ¹³C NMR (126 MHz,DMSO-d₆) δ 175.81, 157.93, 154.78, 135.62, 126.22, 121.10, 117.67,111.87, 109.07, 105.32, 58.97, 28.54; ESI-MS m/z 282.3 (M+H)⁺;MALDI-HRMS m/z calcd for C₁₂H₆ClFN₃O₂(M)⁺ 281.0367. found 281.0850.

Example 5 Exemplary Preparations of Compound 1

To a solution of compound 7 (1 mmol) in 2 mL DMSO, the alkylationreagent (L-R⁶, 1.5 equiv) and potassium carbonate (1.5 equiv) was added.The reaction mixture was allowed to stir at room temperature for 2-2.5 huntil TLC detected the reaction was complete (eluting withDCM:MeOH=10:1). After quenching the reaction with 5 mL water, thereaction mixture was extracted with ethylacetate (5 mL×3). The combinedorganic phases were dried over anhydrous Na₂SO₄. After filtrating andconcentrating in vacuum, compound 1 was obtained by columnchromatography.

Table 4 provides exemplary compounds and conditions for preparation ofcompounds of structure (1) according to the above general procedure.

TABLE 4 Exemplary Preparations of Compound (1) entry 7 8 (L—R⁶) 1 yield%  1 7a CH₃I

  1aa 80  2 7b CH₃I

  1ba 82  3 7c CH₃I

  1ca 81  4 7d CH₃I

  1da 74  5 7e CH₃I

  1ea 78  6 7f CH₃I

  1fa 75  7 7g CH₃I

  1ga 75  8 7h CH₃I

  1ha 78  9 7i CH₃I

  1ia 74 10 7a

  1ab 82 11 7a

  1ac 80 12 7a

  1ad 61 13 7a

  1ae 45 14 7a

  1af 65 15 7a

  1ag 48 16 7a

  1ah 62 17 7a

  1ai 55 18 7a

  1aj 50 19 7a

  1ak 65 20 7g

  1gg 45 21 7a

  1al 77 22 7a

  1am 70 23 7a

  1an 74 24 7a

  1ao 80 25 7a

  1ap 76 26 7a

  1aq 69 27 7a

  1ar 70 28 7a

  1as 63 29 7a

  1at 60 30 7a

  1au 63 31 7a

  1av 71 32 7a

  1aw 65 33 7a

  1ax 75 34 7a

  1ay 78 35 7a

  1az 78 36 7a

  1aaa 35 37 7g

  1gl 75 38 7g

  1gm 71 39 7g

  1gn 74 40 7g

  1go 78 41 7g

  1gp 74 42 7g

  1gp 75 43 7g

  1gu 68 44 7g

  1gv 72

Compounds of structure 1 from Table 4 were analyzed by NMR and massspectrometry. Data is provided below:

1aa: ¹H NMR (500 MHz, DMSO-d₆) δ 11.29 (s, 1H), 8.20 (s, 1H), 7.54 (d,J=7.9 Hz, 1H), 7.24 (s, 1H), 7.15 (d, J=7.5 Hz, 1H), 6.99 (t, J=7.7 Hz,1H), 4.38 (t, J=4.8 Hz, 1H), 3.17 (dd, J=14.9, 4.3 Hz, 1H), 3.10 (dd,J=14.9, 5.8 Hz, 1H), 2.67 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.56,157.43, 133.20, 129.85, 125.96, 120.93, 119.86, 118.12, 116.22, 109.99,57.56, 27.20, 24.30; ESI-MS m/z 300.1 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₃H₁₂ClN₃NaO₂ ⁺ (M+Na)⁺ 300.0510. found 300.0520.1ba: ¹H NMR (500 MHz, DMSO-d₆) δ 10.86 (s, 1H), 8.22 (s, 1H), 7.54 (dd,J=17.0, 7.5 Hz, 1H), 7.45 (d, J=7.4 Hz, 1H), 7.17 (d, J=2.5 Hz, 1H),6.79 (t, J=7.7 Hz, 1H), 4.36 (t, J=5.1 Hz, 1H), 3.10 (dd, J=14.9, 4.6Hz, 1H), 3.04 (dd, J=14.9, 5.6 Hz, 1H), 2.63 (s, 3H); ¹³C NMR (126 MHz,DMSO-d₆) δ 174.56, 157.34, 138.05, 130.29, 128.43, 125.69, 120.80,119.18, 109.99, 77.36, 57.48, 27.21, 24.31; ESI-MS m/z 392.2 (M+Na)⁺;ESI-HRMS m/z calcd for C1₃H1₂IN₃NaO₂ (M+Na)⁺ 391.9866. found 391.9861.1ca: ¹H NMR (400 MHz, DMSO-d₆) δ 10.85 (s, 1H), 8.20 (s, 1H), 7.30 (s,1H), 7.21 (d, J=8.2 Hz, 1H), 7.06 (d, J=2.1 Hz, 1H), 6.87 (d, J=8.2 Hz,1H), 4.32 (t, J=4.8 Hz, 1H), 3.11 (dd, J=14.9, 4.4 Hz, 1H), 3.00 (dd,J=14.9, 5.9 Hz, 1H), 2.67 (s, 3H), 2.37 (s, 3H); ¹³C NMR (126 MHz,DMSO-d₆) δ 174.68, 157.40, 134.76, 128.06, 127.06, 124.50, 122.90,118.43, 111.47, 107.92, 57.68, 27.26, 24.30, 21.79; ESI-MS m/z 280.3(M+Na)⁺; ESI-HRMS m/z calcd for C₁₄H₁₅N₃NaO₂ (M+Na)⁺ 280.1056. found280.1060.1da: ¹H NMR (400 MHz, DMSO-d₆) δ 11.48 (s, 1H), 8.18 (s, 1H), 7.52 (d,J=8.5 Hz, 1H), 7.23 (d, J=2.5 Hz, 1H), 7.18 (d, J=8.5 Hz, 1H), 4.37 (t,J=5.7 Hz, 1H), 3.11 (dd, J=15.2, 4.8 Hz, 1H), 3.05 (dd, J=15.0, 5.7 Hz,1H), 2.63 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.46, 157.34, 134.06,128.28, 126.89, 123.94, 120.62, 119.04, 114.35, 110.39, 57.45, 26.92,24.29; ESI-MS m/z 334.2 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₃H₁₁C₁₂N₃NaO₂(M+Na)⁺ 334.0121. found 334.0108.1ea: ¹H NMR (500 MHz, DMSO-d₆) δ 11.52 (s, 1H), 8.17 (s, 1H), 7.59 (d,J=1.8 Hz, 1H), 7.25 (s, 1H), 7.22 (d, J=1.8 Hz, 1H), 4.36 (t, J=5.1 Hz,1H), 3.09 (dd, J=15.2, 4.9 Hz, 1H), 3.05 (dd, J=15.1, 5.6 Hz, 1H), 2.64(s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.48, 157.32, 131.91, 130.31,127.70, 123.60, 120.50, 117.78, 116.87, 110.02, 57.49, 26.83, 24.28;ESI-MS m/z 334.2 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₃H₁₁C₁₂N₃NaO₂ (M+Na)⁺334.0121. found 334.0130.1fa: ¹H NMR (400 MHz, DMSO-d₆) δ 11.69 (s, 1H), 8.18 (s, 1H), 7.34 (d,J=2.5 Hz, 1H), 7.16 (d, J=8.1 Hz, 1H), 7.04 (d, J=8.1 Hz, 1H), 4.35 (t,J=4.9 Hz, 1H), 3.50 (dd, J=14.8, 4.0 Hz, 1H), 3.04 (dd, J=14.8, 8.9 Hz,1H), 2.81 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.26, 157.50, 134.70,127.93, 125.32, 124.04, 121.54, 120.65, 115.61, 110.99, 58.23, 28.97,24.44; ESI-MS m/z 334.2 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₃H₁₁C₁₂N₃NaO₂(M+Na)⁺ 334.0121. found 334.0109.1ga: ¹H NMR (500 MHz, DMSO-d₆) δ 11.48 (s, 1H), 8.22 (d, J=10.6 Hz, 1H),7.50 (dd, J=8.7, 4.7 Hz, 1H), 7.20 (d, J=2.4 Hz, 1H), 7.02 (dd, J=10.1,8.7 Hz, 1H), 4.35 (t, J=5.6 Hz, 1H), 3.11 (dd, J=15.0, 4.6 Hz, 1H), 3.05(dd, J=14.9, 5.6 Hz, 1H), 2.62 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ174.52, 157.33, 155.10, 153.22, 133.52, 126.45, 118.66, 110.08, 108.33,102.45, 57.44, 26.89, 24.27; ESI-MS m/z 318.1 (M+Na)⁺; ESI-HRMS m/zcalcd for C₁₃H₁₁ClFN₃NaO₂ (M+318.0416. found 318.0428. Chiral HPLC: 98.9enantiomeric excess.1ha: ¹H NMR (400 MHz, DMSO-d₆) δ 11.41 (s, 1H), 8.18 (s, 1H), 7.34 (dd,J=9.8, 2.1 Hz, 1H), 7.26 (d, J=2.2 Hz, 1H), 7.14 (dd, J=9.2, 2.1 Hz,1H), 4.38 (t, J=4.7 Hz, 1H), 3.09 (dd, J=14.6, 4.2 Hz, 1H), 3.05 (dd,J=14.7, 5.1 Hz, 1H), 2.64 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.50,157.35, 155.49, 130.18, 129.27, 127.95, 116.18, 110.35-109.73,103.44-57.48, 26.93, 24.27; ESI-MS m/z 318.2 (M+Na)⁺; ESI-HRMS m/z calcdfor C₁₃H₁₁ClFN₃NaO₂ (M+318.0416. found 318.0418.

1ia: ¹H NMR (400 MHz, DMSO-d₆) δ 11.63 (s, 1H), 8.21 (s, 1H), 7.24 (d,J=2.2 Hz, 1H), 7.12 (dd, J=8.3, 4.1 Hz, 1H), 6.78 (dd, J=10.9, 8.3 Hz,1H), 4.35-4.27 (m, 1H), 3.26 (dd, J=14.7, 4.2 Hz, 1H), 3.00 (dd, J=14.8,8.0 Hz, 1H), 2.76 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.21, 157.47,154.74, 135.66, 126.28, 121.12, 111.85, 109.05, 105.30, 105.13, 57.85,28.71, 24.40; ESI-MS m/z 318.2 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₃H₁₁ClFN₃NaO₂ (M+Na)⁺ 318.0416. found 318.0420.

1ab: ¹H NMR (400 MHz, DMSO-d₆) δ 11.29 (s, 1H), 8.26 (s, 1H), 7.54 (d,J=7.9 Hz, 1H), 7.21 (d, J=2.4 Hz, 1H), 7.14 (d, J=7.4 Hz, 1H), 6.99 (t,J=7.8 Hz, 1H), 5.40 (ddt, J=17.2, 10.1, 4.9 Hz, 1H), 4.78 (dd, J=10.4,1.3 Hz, 1H), 4.50 (dd, J=17.2, 1.3 Hz, 1H), 4.43 (t, J=4.6 Hz, 1H),3.83-3.67 (m, 2H), 3.16 (dd, J=13.5, 3.3 Hz, 1H), 3.11 (dd, J=16.3, 4.5Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.00, 156.80, 133.15, 132.27,132.20, 129.91, 126.15, 126.10, 120.85, 119.83, 118.32, 116.10, 115.91,109.53, 26.95; ESI-MS m/z 326.3 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₆H₁₄ClN₃NaO₂ (M+Na)⁺ 326.06767. found 326.0676.1ac: ¹H NMR (400 MHz, DMSO-d₆) δ 11.27 (s, 1H), 8.10 (s, 1H), 7.51 (d,J=7.9 Hz, 1H), 7.18 (d, J=2.4 Hz, 1H), 7.12 (d, J=7.4 Hz, 1H), 6.97 (t,J=7.8 Hz, 1H), 4.26 (t, J=4.3 Hz, 1H), 3.88-3.78 (m, 1H), 3.13 (dd,J=14.8, 4.5 Hz, 1H), 3.05 (dd, J=14.8, 4.3 Hz, 1H), 0.96 (d, J=6.9 Hz,3H), 0.89 (d, J=6.9 Hz, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.41,157.06, 133.04, 129.88, 126.21, 120.80, 119.77, 118.40, 115.95, 109.11,56.51, 42.27, 26.91, 19.46; ESI-MS m/z 328.3 (M+Na)⁺; ESI-HRMS m/z calcdfor C₁₆H₁₆ClN₃NaO₂(M+Na)⁺ 328.0823. found 328.0829.1ad: ¹H NMR (400 MHz, DMSO-d₆) δ 11.25 (s, 1H), 8.18 (s, 1H), 7.51 (d,J=7.9 Hz, 1H), 7.18 (d, J=2.4 Hz, 1H), 7.11 (d, J=7.1 Hz, 1H), 6.96 (t,J=7.8 Hz, 1H), 4.34 (t, J=4.2 Hz, 1H), 3.17 (dd, J=14.8, 4.4 Hz, 1H),3.11-3.01 (m, 2H), 2.94 (dd, J=13.4, 8.3 Hz, 1H), 1.57 (dt, J=14.9, 7.5Hz, 1H), 1.41-1.28 (m, 2H), 1.20 (tdd, J=12.2, 9.4, 3.8 Hz, 3H),0.88-0.62 (m, 3H): ¹³C NMR (126 MHz, DMSO-d₆) δ 174.45, 157.44, 133.12,129.85, 126.30, 120.73, 119.74, 118.39, 116.04, 109.07, 57.06, 42.21,38.61, 29.59, 26.63, 24.56: ESI-MS m/z 368.3 (M+Na)⁺; ESI-HRMS m/z calcdfor C₁₈H₂₀ClN₃NaO₂(M+Na)⁺ 368.1136. found 368.1145.1ae: ¹H NMR (400 MHz, DMSO-d₆) δ 11.32 (s, 1H), 8.24 (s, 1H), 7.58 (dd,J=7.8, 4.8 Hz, 1H), 7.25 (d, J=2.3 Hz, 1H), 7.18 (dd, J=7.3, 4.0 Hz,1H), 7.03 (dd, J=7.8, 3.7 Hz, 1H), 5.40 (ddt, J=17.5, 10.9, 6.8 Hz, 1H),4.84 (dd, J=13.0, 6.1 Hz, 2H), 4.41 (t, J=4.8 Hz, 1H), 3.32-3.03 (m,4H), 1.88 (tt, J=14.4, 7.1 Hz, 2H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.53,157.36, 135.02, 133.12, 129.82, 126.17, 120.82, 119.84, 118.28, 116.91,116.10, 109.27, 57.18, 37.09, 31.78, 26.83; ESI-MS m/z 340.3 (M+Na)⁺;ESI-HRMS m/z calcd for C₁₆H₁₆ClN₃NaO₂ (M+Na)⁺ 340.0823. found 340.0811.1af: ¹H NMR (400 MHz, DMSO-d₆) δ 11.29 (s, 1H), 8.19 (s, 1H), 7.53 (d,J=7.9 Hz, 1H), 7.26-7.19 (m, 3H), 7.14 (dd, J=7.3, 4.5 Hz, 2H), 7.05 (d,J=7.1 Hz, 2H), 6.98 (t, J=7.8 Hz, 1H), 4.36 (t, J=4.4 Hz, 1H), 3.23-3.02(m, 4H), 2.31 (dt, J=10.0, 6.6 Hz, 2H), 1.13-0.96 (m, 4H); ¹³C NMR (126MHz, DMSO-d₆) δ 174.40, 157.30, 142.31, 133.14, 129.89, 128.65, 126.27,126.21, 126.02, 120.85, 119.80, 118.27, 116.14, 109.31, 57.16, 37.46,34.90, 28.07, 27.28, 26.90; ESI-MS m/z 418.4 (M+Na)⁺; ESI-HRMS m/z calcdfor C₂₂H₂₂ClN₃NaO₂ (M+Na)⁺ 418.1293. found 418.1284.1ag (i.e., “la”): ¹H NMR (400 MHz, DMSO-d₆) δ 11.28 (s, 1H), 8.22 (d,J=4.0 Hz, 1H), 7.51 (dd, J=7.9, 3.6 Hz, 1H), 7.19 (d, J=2.2 Hz, 1H),7.13 (d, J=7.4 Hz, 1H), 6.98 (dd, J=7.8, 2.4 Hz, 1H), 4.40-4.33 (m, 1H),3.67-3.51 (m, 2H), 3.51-3.39 (m, 1H), 3.31-2.93 (m, 5H), 1.60 (ddd,J=37.4, 21.3, 6.9 Hz, 2H), 1.00 (dd, J=13.4, 5.5 Hz, 1H); ¹³C NMR (126MHz, DMSO-d₆) δ 174.54, 157.18, 133.11, 129.76, 126.34, 120.82, 119.87,118.28, 116.07, 109.00, 75.23, 67.36, 57.09, 41.35, 28.54, 26.73, 24.92;ESI-MS m/z 370.3 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₇H₁₈ClN₃NaO₃ (M+Na)⁺370.0929. found 370.0935.1ah: ¹H NMR (400 MHz, DMSO-d₆) δ 11.26 (s, 1H), 8.25 (s, 1H), 7.53 (d,J=7.9 Hz, 1H), 7.17 (d, J=2.4 Hz, 1H), 7.12 (d, J=7.2 Hz, 1H), 6.98 (t,J=7.8 Hz, 1H), 4.36 (t, J=3.8 Hz, 1H), 3.55 (dd, J=13.4, 2.4 Hz, 1H),3.49 (dd, J=11.4, 2.5 Hz, 1H), 3.21 (dd, J=14.8, 3.9 Hz, 1H), 3.04 (dd,J=11.4, 3.4 Hz, 1H), 2.99 (t, J=8.4 Hz, 1H), 2.85 (ddd, J=14.2, 7.8, 3.2Hz, 2H), 2.70 (ddd, J=11.7, 10.1, 1.9 Hz, 1H), 1.15 (ddd, J=10.8, 9.2,5.5 Hz, 1H), 0.81-0.60 (m, 3H), 0.39 (d, J=11.6 Hz, 1H); ¹³C NMR (126MHz, DMSO-d₆) δ 174.25, 157.30, 132.96, 129.88, 126.39, 120.79, 119.82,118.48, 116.04, 108.99, 66.77, 57.14, 43.22, 33.61, 30.02, 26.47; ESI-MSm/z 384.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₈H₂₀ClN₃NaO₃(M+Na)⁺384.1085. found 384.1096.1ai: ¹H NMR (400 MHz, DMSO-d₆) δ 11.29 (s, 1H), 8.28 (s, 1H), 7.52 (d,J=7.9 Hz, 1H), 7.20 (dd, J=5.9, 2.4 Hz, 1H), 7.14 (d, J=7.6 Hz, 1H),6.99 (t, J=7.8 Hz, 1H), 4.42 (t, J=4.8 Hz, 1H), 3.47-3.38 (m, 1H), 3.20(ddd, J=27.7, 14.4, 5.0 Hz, 1H), 3.12 (d, J=4.6 Hz, 2H), 2.69-2.59 (m,1H), 2.41-2.34 (m, 1H), 2.15 (ddd, J=12.9, 5.0, 2.5 Hz, 1H); ¹³C NMR(126 MHz, DMSO-d₆) δ 174.27, 156.81, 133.14, 129.87, 126.07, 120.86,119.84, 118.21, 116.09, 109.47, 57.44, 48.70, 48.50, 45.21, 27.04;ESI-MS m/z 342.2 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₆H₁₄ClN₃NaO₃(M+Na)⁺342.0616. found 342.0611.1aj: ¹H NMR (400 MHz, DMSO-d₆) δ 11.28 (s, 1H), 8.26 (s, 1H), 7.54 (d,J=7.9 Hz, 1H), 7.18 (d, J=2.2 Hz, 1H), 7.12 (d, J=7.4 Hz, 1H), 6.98 (t,J=7.8 Hz, 1H), 4.36 (t, J=3.8 Hz, 1H), 3.63 (s, 2H), 3.22 (dd, J=14.8,3.8 Hz, 1H), 3.10-2.94 (m, 2H), 2.84 (dd, J=13.5, 8.4 Hz, 1H), 2.26 (s,1H), 2.16-2.04 (m, 1H), 1.36 (s, 9H), 1.11 (s, 1H), 0.49 (s, 4H); ¹³CNMR (126 MHz, DMSO-d₆) δ 174.23, 157.27, 154.12, 132.95, 129.89, 126.40,120.79, 119.83, 118.48, 116.08, 108.97, 78.80, 57.13, 42.95, 34.50,29.16, 29.00, 28.52, 26.44; ESI-MS m/z 483.5 (M+Na)⁺; ESI-HRMS m/z calcdfor C₂₃H₂₉ClN₄NaO₄ (M+Na)⁺ 483.1769. found 483.1786.1ak: ¹H NMR (500 MHz, DMSO-d₆) δ 11.31 (s, 1H), 8.19 (s, 1H), 7.52 (d,J=7.9 Hz, 1H), 7.24 (dd, J=8.4, 7.5 Hz, 2H), 7.19 (d, J=2.3 Hz, 1H),7.13 (d, J=7.5 Hz, 1H), 6.98 (t, J=7.8 Hz, 1H), 6.89 (t, J=7.3 Hz, 1H),6.73 (d, J=8.0 Hz, 2H), 4.35 (t, J=4.5 Hz, 1H), 3.47-3.38 (m, 1H),3.32-3.26 (m, 3H), 3.15 (dd, J=14.8, 4.7 Hz, 1H), 3.08 (dd, J=14.9, 4.4Hz, 1H), 1.52 (dd, J=12.4, 6.7 Hz, 1H), 1.42 (dd, J=12.2, 6.7 Hz, 1H);¹³C NMR (126 MHz, DMSO-d₆) δ 174.48, 158.83, 157.17, 133.12, 129.83,129.80, 126.27, 120.90, 120.82, 119.87, 118.33, 116.17, 114.70, 109.35,64.54, 57.17, 34.97, 27.59, 26.98; ESI-MS m/z 420.4 (M+Na)⁺; ESI-HRMSm/z calcd for C₂₁H₂₀ClN₃NaO₃ (M+Na)⁺ 420.1085. found 420.1066.1al: ¹H NMR (400 MHz, DMSO-d₆) δ 11.35 (s, 1H), 8.44 (s, 1H), 7.57 (d,J=7.9 Hz, 1H), 7.22 (d, J=2.5 Hz, 1H), 7.18 (d, J=7.8 Hz, 1H), 7.15 (d,J=7.3 Hz, 1H), 7.06 (t, J=5.7 Hz, 1H), 6.97 (t, J=7.8 Hz, 1H), 6.72 (dd,J=7.6, 0.9 Hz, 1H), 5.86 (dd, J=7.8, 1.2 Hz, 1H), 4.52 (t, J=4.1 Hz,1H), 4.41 (d, J=16.2 Hz, 1H), 4.33 (d, J=16.2 Hz, 1H), 3.25 (dd, J=14.9,4.5 Hz, 1H), 3.13 (dd, J=14.9, 4.2 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ174.04, 160.80, 158.85, 156.64, 133.13, 129.95, 129.26, 127.76, 126.36,124.38, 123.46, 120.93, 119.97, 118.43, 116.16, 115.41, 109.28, 57.75,26.53; ESI-MS m/z 394.3 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₅ClFN₃NaO₂(M+Na)⁺ 394.0729. found 394.0735.1am: ¹H NMR (400 MHz, DMSO-d₆) δ 11.33 (s, 1H), 8.44 (s, 1H), 7.59 (d,J=7.9 Hz, 1H), 7.23 (d, J=2.4 Hz, 1H), 7.18 (d, J=7.4 Hz, 1H), 7.09 (dd,J=14.3, 8.1 Hz, 1H), 7.01 (dd, J=13.4, 5.6 Hz, 2H), 6.71 (d, J=9.9 Hz,1H), 6.25 (d, J=7.6 Hz, 1H), 4.56 (t, J=4.4 Hz, 1H), 4.45 (d, J=15.9 Hz,1H), 4.32 (d, J=15.9 Hz, 1H), 3.26 (dd, J=14.9, 4.6 Hz, 1H), 3.16 (dd,J=14.9, 4.3 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.10, 163.31,161.37, 156.79, 139.61, 133.11, 130.42, 129.87, 126.24, 122.36, 120.88,119.88, 118.31, 116.21, 114.25, 113.93, 109.99, 57.67, 26.60; ESI-MS m/z394.3 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₅ClFN₃NaO₂ (M+Na)⁺ 394.0729.found 394.0722.1an: ¹H NMR (400 MHz, DMSO-d₆) δ 11.35 (s, 1H), 8.51 (s, 1H), 7.55 (d,J=7.9 Hz, 1H), 7.22 (d, J=3.5 Hz, 1H), 7.20 (d, J=11.1 Hz, 1H), 7.14 (d,J=7.5 Hz, 1H), 6.96 (t, J=7.8 Hz, 1H), 6.70 (ddd, J=13.1, 6.5, 1.3 Hz,1H), 5.70 (t, J=7.1 Hz, 1H), 4.52 (t, J=4.2 Hz, 1H), 4.44 (d, J=16.2 Hz,1H), 4.35 (d, J=16.1 Hz, 1H), 3.24 (dd, J=14.9, 4.4 Hz, 1H), 3.12 (dd,J=14.9, 4.2 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 173.96, 156.48,150.79, 148.74, 146.67, 133.08, 129.88, 126.40, 126.04, 124.56, 123.05,120.89, 119.92, 118.41, 116.51, 116.12, 109.16, 57.71, 26.48; ESI-MS m/z412.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₄ClF₂N₃NaO₂ (M+Na)⁺ 412.0635.found 412.0642.1ao: ¹H NMR (400 MHz, DMSO-d₆) δ 11.27 (s, 1H), 8.42 (s, 1H), 7.52 (d,J=7.9 Hz, 1H), 7.17 (d, J=2.4 Hz, 1H), 7.13 (d, J=7.4 Hz, 1H), 6.99 (dd,J=8.5, 2.3 Hz, 1H), 6.95 (dd, J=9.3, 6.2 Hz, 1H), 6.83 (ddd, J=11.2,7.8, 2.0 Hz, 1H), 6.24-6.14 (m, 1H), 4.49 (t, J=4.1 Hz, 1H), 4.35 (d,J=15.7 Hz, 1H), 4.23 (d, J=15.7 Hz, 1H), 3.22 (dd, J=14.9, 4.4 Hz, 1H),3.10 (dd, J=14.9, 4.3 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.03,156.73, 150.27, 148.31, 134.43, 133.03, 129.80, 126.27, 123.28, 120.84,119.84, 118.32, 117.34, 117.21, 116.34, 116.16, 109.10, 57.59, 26.51;ESI-MS m/z 412.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₄ClF₂N₃NaO₂(M+Na)⁺ 412.0635. found 412.0635.1ap: ¹H NMR (400 MHz, DMSO-d₆) δ 11.26 (s, J=1.4 Hz, 1H), 8.50 (s, 1H),7.53 (d, J=7.9 Hz, 1H), 7.20 (d, J=2.4 Hz, 1H), 7.11 (d, J=7.4 Hz, 1H),7.03-6.93 (m, 2H), 6.41 (d, J=6.2 Hz, 2H), 4.53 (t, J=4.3 Hz, 1H), 4.42(d, J=16.0 Hz, 1H), 4.31 (d, J=16.0 Hz, 1H), 3.22 (dd, J=15.0, 4.6 Hz,1H), 3.14 (dd, J=14.9, 4.4 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.15,163.60, 161.63, 156.71, 141.34, 133.09, 129.75, 126.20, 120.86, 119.75,118.13, 116.38, 110.11, 109.95, 109.22, 103.14, 102.94, 57.72, 26.61;ESI-MS m/z 412.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₆H₁₄ClF₂N₃NaO₂(M+Na)⁺ 412.0635. found 412.0628.1aq: ¹H NMR (500 MHz, DMSO-d₆) δ 11.28 (s, 1H), 8.46 (s, 1H), 7.48 (d,J=7.9 Hz, 1H), 7.14 (dd, J=27.0, 4.9 Hz, 2H), 6.92 (t, J=7.8 Hz, 1H),6.73 (dd, J=9.5, 1.9 Hz, 1H), 5.88 (dd, J=8.2, 2.2 Hz, 1H), 4.47 (t,J=4.2 Hz, 1H), 4.36 (d, J=15.8 Hz, 1H), 4.28 (d, J=15.8 Hz, 1H), 3.21(dd, J=14.9, 4.3 Hz, 1H), 3.08 (dd, J=14.9, 4.2 Hz, 1H); ¹³C NMR (500MHz, DMSO-d₆) δ 173.88, 156.44, 139.96, 137.98, 132.99, 129.76, 126.37,122.72, 122.65, 121.62, 120.82, 119.85, 118.36, 116.04, 112.23, 112.06,108.99, 57.56, 26.48; ESI-MS m/z 430.3 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₆H₁₃ClF₃N₃NaO₂ (M+Na)⁺ 430.0540. found 430.0543.1ar: ¹H NMR (500 MHz, DMSO-d₆) δ 11.29 (s, 1H), 8.37 (s, 1H), 7.55 (d,J=7.8 Hz, 1H), 7.18 (d, J=8.4 Hz, 1H), 7.13 (d, J=7.5 Hz, 1H), 6.94 (dd,J=15.3, 7.6 Hz, 2H), 6.72 (s, 1H), 6.24 (d, J=6.4 Hz, 1H), 4.47 (t,J=5.4 Hz, 1H), 4.35 (d, J=15.6 Hz, 1H), 4.21 (d, J=15.6 Hz, 1H), 3.18(dd, J=14.6, 4.3 Hz, 1H), 3.12 (dd, J=14.8, 3.7 Hz, 1H), 2.28 (s, 1H),2.16 (s, 2H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.16, 156.93, 137.70,136.74, 133.12, 129.97, 128.47, 127.97, 127.63, 126.25, 123.64, 120.86,119.86, 118.33, 116.16, 109.41, 63.33, 57.62, 26.62, 21.32; ESI-MS m/z390.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₂₀H₁₈ClN₃NaO₂(M+Na)⁺ 390.0980.found 390.0975.1as: ¹H NMR (500 MHz, DMSO-d₆) δ 11.32 (d, J=1.4 Hz, 1H), 8.44 (s, 1H),7.56 (d, J=7.9 Hz, 1H), 7.21 (d, J=2.4 Hz, 1H), 7.14 (d, J=7.5 Hz, 1H),6.96 (t, J=7.8 Hz, 1H), 6.84 (d, J=7.9 Hz, 2H), 6.43 (d, J=7.9 Hz, 2H),4.46 (t, J=4.2 Hz, 1H), 4.34 (d, J=15.5 Hz, 1H), 4.18 (d, J=15.5 Hz,1H), 3.22 (dd, J=14.9, 4.5 Hz, 1H), 3.13 (dd, J=14.9, 4.3 Hz, 1H), 2.19(s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.10, 156.89, 136.18, 133.72,133.13, 130.00, 129.05, 127.83, 126.37, 120.82, 119.85, 118.42, 116.17,109.24, 57.60, 26.53, 21.17; ESI-MS m/z 390.4 (M+Na)⁺; ESI-HRMS m/zcalcd for C₂₀H₁₈ClN₃NaO₂(M+Na)⁺ 390.0980. found 390.0983.1at: ¹H NMR (400 MHz, DMSO-d₆) δ 11.34 (s, 1H), 8.39 (s, 1H), 7.55 (d,J=7.9 Hz, 1H), 7.35 (d, J=8.3 Hz, 2H), 7.19 (d, J=2.4 Hz, 1H), 7.16 (d,J=7.5 Hz, 1H), 6.96 (t, J=7.8 Hz, 1H), 6.21 (d, J=8.3 Hz, 2H), 4.49 (t,J=4.0 Hz, 1H), 4.35 (d, J=15.9 Hz, 1H), 4.16 (d, J=15.9 Hz, 1H), 3.25(dd, J=14.9, 4.2 Hz, 1H), 3.09 (dd, J=14.9, 4.2 Hz, 1H); ¹³C NMR (126MHz, DMSO-d₆) δ 174.00, 156.73, 137.17, 136.50, 133.07, 129.94, 128.74,126.40, 120.90, 119.97, 118.48, 116.14, 109.15, 93.10, 57.66, 26.40;ESI-MS m/z 502.3 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₅ClIN₃NaO₂ (M+Na)⁺501.9789. found 501.9784.1au: ¹H NMR (400 MHz, DMSO-d₆) δ 11.33 (s, 1H), 8.41 (s, 1H), 7.56 (d,J=7.9 Hz, 1H), 7.20 (d, J=2.4 Hz, 1H), 7.16 (d, J=7.5 Hz, 1H), 6.97 (t,J=7.8 Hz, 1H), 6.83 (d, J=8.9 Hz, 1H), 6.54 (dd, J=8.6, 5.5 Hz, 2H),4.49 (t, J=4.1 Hz, 1H), 4.38 (d, J=15.6 Hz, 1H), 4.22 (d, J=15.6 Hz,1H), 3.24 (dd, J=14.9, 4.3 Hz, 1H), 3.12 (dd, J=14.9, 4.2 Hz, 1H); ¹³CNMR (126 MHz, DMSO-d₆) δ 174.00, 156.73, 137.17, 136.50, 133.07, 129.94,128.74, 126.40, 120.90, 119.97, 118.48, 116.14, 109.15, 93.10, 57.66,26.40; ESI-MS m/z 394.3 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₅ClFN₃NaO₂⁺ (M+Na)⁺ 394.0729. found 394.0730.1av: ¹H NMR (400 MHz, DMSO-d₆) δ 11.29 (s, 1H), 8.42 (s, 1H), 7.55 (d,J=7.9 Hz, 1H), 7.21 (dd, J=11.7, 1.9 Hz, 2H), 7.14 (d, J=7.5 Hz, 1H),7.05 (d, J=2.8 Hz, 1H), 7.02 (d, J=7.9 Hz, 1H), 6.97 (t, J=7.8 Hz, 1H),6.28 (d, J=7.7 Hz, 1H), 4.52 (t, J=4.4 Hz, 1H), 4.40 (d, J=15.8 Hz, 1H),4.27 (d, J=15.8 Hz, 1H), 3.21 (dd, J=13.9, 3.7 Hz, 1H), 3.12 (dd,J=14.9, 4.4 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.12, 156.77,139.26, 133.25, 133.11, 130.32, 129.88, 127.36, 127.19, 126.24, 125.02,120.90, 119.89, 118.31, 116.19, 109.29, 57.67, 40.56, 40.48, 40.31,40.14, 39.98, 39.81, 39.64, 39.48, 26.60; ESI-MS m/z 410.3 (M+Na)⁺;ESI-HRMS m/z calcd for C₁₉H₁₅Cl₂N₃NaO₂(M+Na)⁺ 410.0433. found 410.0433.1aw: ¹H NMR (500 MHz, DMSO-d₆) δ 11.39 (s, 1H), 8.42 (s, 1H), 7.60 (dd,J=5.6, 3.8 Hz, 3H), 7.46 (t, J=7.7 Hz, 2H), 7.35 (t, J=14.7 Hz, 1H),7.30 (d, J=8.2 Hz, 2H), 7.24 (d, J=2.3 Hz, 1H), 7.17 (d, J=7.4 Hz, 1H),6.99 (t, J=7.8 Hz, 1H), 6.49 (d, J=8.1 Hz, 2H), 4.53 (t, J=4.0 Hz, 1H),4.46 (d, J=15.9 Hz, 1H), 4.27 (d, J=15.9 Hz, 1H), 3.28 (dd, J=14.9, 4.2Hz, 1H), 3.14 (dd, J=14.9, 4.1 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ174.10, 156.87, 140.24, 139.08, 135.88, 133.16, 130.09, 129.33, 127.81,126.95, 126.78, 126.47, 120.94, 120.00, 118.58, 116.16, 109.27, 57.74,26.44; ESI-MS m/z 452.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₂₆H₂₀ClN₃NaO₂(M+H)⁺ 452.1136. found 452.1133.1ax: ¹H NMR (400 MHz, DMSO-d₆) δ 11.41 (s, 1H), 8.51 (s, 1H), 7.57 (d,J=7.9 Hz, 1H), 7.33 (dd, J=8.7, 2.6 Hz, 1H), 7.22 (d, J=2.4 Hz, 1H),7.16 (d, J=7.4 Hz, 1H), 6.97 (t, J=7.8 Hz, 1H), 6.59 (dd, J=8.5, 2.6 Hz,1H), 5.62 (dd, J=8.6, 6.1 Hz, 1H), 4.55 (t, J=3.9 Hz, 1H), 4.37 (d,J=16.5 Hz, 1H), 4.28 (d, J=16.6 Hz, 1H), 3.27 (dd, J=14.9, 4.1 Hz, 1H),3.13 (dd, J=14.9, 4.1 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.01,162.06, 160.10, 156.53, 133.07, 132.31, 129.94, 129.87, 128.09, 126.50,121.01, 120.08, 118.61, 116.95, 116.07, 113.96, 109.19, 57.86, 26.35;ESI-MS m/z 428.3 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₆H₁₄Cl₂FN₃NaO₂(M+Na)⁺ 428.0339. found 428.0340.1ay: ¹H NMR (400 MHz, DMSO-d₆) δ 11.35 (s, 1H), 8.50 (s, 1H), 7.59 (d,J=7.2 Hz, 1H), 7.55 (d, J=8.0 Hz, 1H), 7.21 (d, J=2.4 Hz, 1H), 7.14 (d,J=7.2 Hz, 1H), 6.96 (t, J=7.8 Hz, 1H), 6.90 (t, J=7.8 Hz, 1H), 6.16 (t,J=7.2 Hz, 1H), 4.54 (t, J=4.1 Hz, 1H), 4.48 (d, J=16.3 Hz, 1H), 4.38 (d,J=16.2 Hz, 1H), 3.25 (dd, J=14.9, 4.4 Hz, 1H), 3.12 (dd, J=14.9, 4.2 Hz,1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.12, 157.55, 156.53, 155.51,133.04, 132.51, 129.77, 126.35, 125.39, 124.58, 123.99, 121.83, 120.95,120.02, 118.40, 116.13, 109.02, 57.77, 34.37, 26.31; ESI-MS m/z 462.4(M+Na)⁺; ESI-HRMS m/z calcd for C₂₀H₁₄ClF₄N₃NaO₂ (M+Na)⁺ 462.0603. found462.0616.1az: ¹H NMR (400 MHz, DMSO-d₆) δ 11.24 (s, 1H), 8.43 (s, 1H), 7.64 (d,J=12.1 Hz, 2H), 7.48 (d, J=7.9 Hz, 1H), 7.39 (s, 1H), 7.20 (d, J=2.2 Hz,1H), 7.12 (d, J=7.5 Hz, 1H), 6.94 (t, J=7.8 Hz, 1H), 4.59-4.50 (m, 3H),3.16 (dd, J=14.9, 4.6 Hz, 1H), 3.10 (dd, J=14.9, 5.8 Hz, 1H); ¹³C NMR(126 MHz, DMSO-d₆) δ 174.32, 156.67, 136.63, 135.07, 133.17, 130.84,129.68, 128.32, 128.06, 126.16, 126.02, 125.04, 122.87, 120.88, 119.80,117.96, 116.21, 109.41, 57.68, 26.92; ESI-MS m/z 478.4 (M+Na)⁺; ESI-HRMSm/z calcd for C₂₀H₁₄Cl₂F₃N₃NaO₂ (M+Na)⁺ 478.0307. found 478.0326.1aaa: ¹H NMR (500 MHz, DMSO-d₆) δ 11.39 (s, 1H), 8.46 (s, 1H), 8.16 (dd,J=4.5, 1.5 Hz, 2H), 7.56 (d, J=7.9 Hz, 1H), 7.20 (d, J=2.4 Hz, 1H), 7.16(d, J=7.3 Hz, 1H), 6.96 (t, J=7.8 Hz, 1H), 6.29 (d, J=5.8 Hz, 2H), 4.54(t, J=4.0 Hz, 1H), 4.42 (d, J=16.7 Hz, 1H), 4.24 (d, J=16.7 Hz, 1H),3.27 (dd, J=14.9, 4.1 Hz, 1H), 3.11 (dd, J=14.9, 4.2 Hz, 1H); ¹³C NMR(126 MHz, DMSO-d₆) δ 174.01, 156.61, 149.60, 145.62, 133.11, 129.97,126.51, 126.45, 121.31, 121.19, 121.02, 120.05, 118.59, 118.51, 116.15,109.19, 57.84, 26.37; ESI-MS m/z 377.3 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₈H₁₅ClN₄NaO₂ (M+Na)⁺ 377.0776. found 377.0791.1gaa: ¹H NMR (400 MHz, DMSO-d₆) δ 11.53 (s, 1H), 8.44 (s, 1H), 8.15 (d,J=5.7 Hz, 2H), 7.51 (dd, J=8.6, 4.9 Hz, 1H), 7.18 (s, 1H), 6.97 (t,J=9.4 Hz, 1H), 6.31 (d, J=5.4 Hz, 2H), 4.51 (s, 1H), 4.39 (d, J=16.7 Hz,1H), 4.23 (d, J=16.5 Hz, 1H), 3.23 (dd, J=14.8, 4.0 Hz, 1H), 3.07 (dd,J=15.1, 4.0 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 173.98, 156.61,155.13, 153.25, 149.56, 145.60, 133.48, 126.90, 126.03, 121.35, 119.06,109.43, 108.57, 102.41, 57.74, 26.19; ESI-MS m/z 373.2 (M+H)⁺; ESI-HRMSm/z calcd for C₁₈H₁₆ClN₄NaO₂ ⁺ (M+H)⁺373.0861. found 373.0873.1gl: ¹H NMR (500 MHz, DMSO-d₆) δ 11.54 (s, 1H), 8.51 (s, 1H), 7.52 (dd,J=8.7, 4.7 Hz, 1H), 7.22 (d, J=2.3 Hz, 1H), 7.18 (dd, J=7.4, 7.4 Hz,1H), 7.04 (dd, J=14.1, 4.5 Hz, 1H), 6.97 (dd, J=10.1, 8.7 Hz, 1H), 6.70(t, J=7.5 Hz, 1H), 5.86 (t, J=7.5 Hz, 1H), 4.50 (t, J=4.2 Hz, 1H), 4.38(d, J=16.2 Hz, 1H), 4.29 (d, J=16.1 Hz, 1H), 3.22 (dd, J=14.9, 4.3 Hz,1H), 3.11 (dd, J=14.9, 4.3 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ174.01,160.78, 158.84, 156.60, 155.10, 153.22, 133.48, 129.28, 127.74, 126.01,124.21, 123.44, 118.99, 115.43, 109.40, 108.44, 102.40, 57.64, 26.26;ESI-MS m/z 412.3 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₄ClF₂N₃NaO₂(M+Na)⁺ 412.0635. found 412.0642.1gm: ¹H NMR (500 MHz, DMSO-d₆) δ 11.44 (s, 1H), 8.45 (s, 1H), 7.49 (dd,J=8.7, 4.7 Hz, 1H), 7.19 (d, J=2.4 Hz, 1H), 7.03 (dd, J=7.9, 6.2 Hz,1H), 6.95 (ddd, J=7.7, 6.3, 5.4 Hz, 2H), 6.54 (d, J=8.3 Hz, 1H), 6.25(d, J=7.7 Hz, 1H), 4.49 (td, J=4.2, 0.7 Hz, 1H), 4.38 (d, J=15.9 Hz,1H), 4.25 (d, J=15.9 Hz, 1H), 3.20 (dd, J=14.9, 4.4 Hz, 1H), 3.09 (dd,J=14.9, 4.3 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ174.07, 163.27, 161.33,156.78, 155.10, 153.22, 139.56, 133.47, 130.28, 126.78, 125.90, 122.46,118.82, 114.22, 109.33, 108.35, 102.51, 57.56, 26.31; ESI-MS m/z 412.3(M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₄ClF₂N₃NaO₂ (M+Na)⁺ 412.0635. found412.0641.1gn: ¹H NMR (500 MHz, DMSO-d₆) δ 11.48 (s, 1H), 8.48 (s, 1H), 7.49 (dd,J=8.7, 4.7 Hz, 1H), 7.25-7.15 (m, 2H), 6.96 (dd, J=10.1, 8.7 Hz, 1H),6.70 (dd, J=8.1, 1.5 Hz, 2H), 5.78 (t, J=7.1 Hz, 1H), 4.49 (t, J=4.1 Hz,1H), 4.41 (d, J=16.0 Hz, 1H), 4.32 (d, J=16.0 Hz, 1H), 3.21 (dd, J=14.9,4.3 Hz, 1H), 3.08 (dd, J=14.9, 4.2 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆)173.95, 156.50, 155.09, 153.20, 150.77, 148.65, 146.68, 133.44, 126.85,125.88, 124.30, 123.16, 118.85, 116.33, 109.22, 108.36, 102.40, 57.61,26.20; ESI-MS m/z 430.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₃ClF₃N₃NaO₂(M+Na)⁺ 430.0540. found 430.0545.1go: ¹H NMR (500 MHz, DMSO-d₆) δ 11.40 (s, 1H), 8.41 (s, 1H), 7.46 (dd,J=8.7, 4.7 Hz, 1H), 7.15 (d, J=2.4 Hz, 1H), 7.00 (dd, J=8.5, 2.2 Hz,1H), 6.94 (dd, J=10.0, 8.8 Hz, 1H), 6.71 (ddd, J=11.1, 7.8, 2.0 Hz, 1H),6.31 (s, 1H), 4.47 (t, J=4.2 Hz, 1H), 4.32 (d, J=15.6 Hz, 1H), 4.21 (d,J=15.6 Hz, 1H), 3.20 (dd, J=14.9, 4.2 Hz, 1H), 3.06 (dd, J=14.9, 4.3 Hz,1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 173.99, 156.76, 155.10, 153.22,150.35, 148.39, 134.38, 133.40, 126.74, 125.80, 123.50, 118.80, 117.26,116.29, 109.19, 108.14, 102.47, 57.58, 26.30; ESI-MS m/z 430.4 (M+Na)⁺;ESI-HRMS m/z calcd for C₁₉H₁₃ClF₃N₃NaO₂ (M+Na)⁺ 430.0540. found430.0552.1gp: ¹H NMR (400 MHz, DMSO-d₆) δ 11.38 (s, 1H), 8.43 (s, 1H), 7.47 (dd,J=8.7, 4.7 Hz, 1H), 7.17 (d, J=2.4 Hz, 1H), 7.00-6.92 (m, 2H), 6.33 (dd,J=8.2, 2.1 Hz, 2H), 4.51 (t, J=4.1 Hz, 1H), 4.40 (d, J=16.0 Hz, 1H),4.28 (d, J=16.0 Hz, 1H), 3.21 (dd, J=14.9, 4.3 Hz, 1H), 3.08 (dd,J=14.9, 4.3 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.12, 163.47,161.51, 156.77, 155.10, 153.21, 141.11, 133.43, 126.71, 125.68, 118.55,110.03, 109.83, 108.28, 102.69, 57.57, 26.17; ESI-MS m/z 430.4 (M+Na)⁺;ESI-HRMS m/z calcd for C₁₉H₁₃ClF₃N₃NaO₂ (M+Na)⁺ 430.0540. found430.0541.1gq: ¹H NMR (400 MHz, DMSO-d₆) δ 11.43 (s, 1H), 8.43 (s, 1H), 7.44 (dd,J=8.7, 4.7 Hz, 1H), 7.16 (d, J=2.4 Hz, 1H), 6.95 (dd, J=10.1, 8.7 Hz,1H), 6.81 (dd, J=6.4, 4.9 Hz, 1H), 6.08 (dd, J=10.6, 2.4 Hz, 1H), 4.47(t, J=4.0 Hz, 1H), 4.37 (d, J=15.6 Hz, 1H), 4.29 (d, J=15.6 Hz, 1H),3.20 (dd, J=14.9, 4.2 Hz, 1H), 3.05 (dd, J=14.9, 4.2 Hz, 1H); ¹³C NMR(126 MHz, DMSO-d₆) δ 173.83, 156.49, 155.07, 153.18, 139.92, 137.94,133.35, 126.74, 125.71, 123.01, 121.43, 118.75, 111.88, 109.05, 108.24,102.31, 57.41, 34.16, 26.25; ESI-MS m/z 448.4 (M+Na)⁺; ESI-HRMS m/zcalcd for C₁₉H₁₂ClF₄N₃NaO₂ (M+Na)⁺ 448.0446. found 448.0430.1gu: ¹H NMR (500 MHz, DMSO-d₆) δ 11.48 (s, 1H), 8.42 (s, 1H), 7.50 (dd,J=8.7, 4.7 Hz, 1H), 7.18 (d, J=2.4 Hz, 1H), 6.97 (dd, J=10.1, 8.7 Hz,1H), 6.82 (t, J=6.1 Hz, 2H), 6.56 (dd, J=8.4, 5.6 Hz, 2H), 4.46 (t,J=3.8 Hz, 1H), 4.34 (d, J=15.6 Hz, 1H), 4.20 (d, J=15.6 Hz, 1H), 3.20(dd, J=14.9, 4.3 Hz, 1H), 3.07 (dd, J=14.9, 4.2 Hz, 1H); ¹³C NMR (126MHz, DMSO-d₆) δ 174.06, 162.39, 160.46, 156.82, 155.09, 153.21, 133.40,132.81, 128.62, 126.80, 125.95, 118.95, 115.12, 114.96, 109.27, 108.39,102.18, 57.52, 26.17; ESI-MS m/z 412.3 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₉H₁₄ClF₂N₃NaO₂ ⁺ (M+Na)⁺ 412.0635. found 412.0643.1gv: ¹H NMR (400 MHz, DMSO-d₆) δ 11.43 (s, 1H), 8.39 (s, 1H), 7.50 (dd,J=8.7, 4.7 Hz, 1H), 7.21 (d, J=9.2 Hz, 1H), 7.18 (d, J=2.3 Hz, 1H), 7.01(dt, J=18.9, 8.9 Hz, 3H), 6.34 (d, J=7.7 Hz, 1H), 4.50 (t, J=4.4 Hz,1H), 4.38 (d, J=15.8 Hz, 1H), 4.26 (d, J=15.8 Hz, 1H), 3.20 (dd, J=14.9,4.5 Hz, 1H), 3.09 (dd, J=14.9, 4.3 Hz, 1H); ¹³C NMR (126 MHz, DMSO-d₆) δ174.11, 156.82, 155.11, 153.23, 139.11, 133.47, 133.25, 130.14, 127.34,127.14, 126.70, 125.87, 125.11, 118.75, 109.31, 108.39, 102.52, 57.58,26.27; ESI-MS m/z 428.4 (M+Na)⁺; ESI-HRMS m/z calcd for C₁₉H₁₄Cl₂FN₃NaO₂(M+Na)⁺ 428.0339. found 428.0327.

Example 6 Exemplary Halogenation

Compounds of structure (III), wherein R⁵ is halogen (i.e., compound 1′″,wherein R⁵ is R^(5′)) were prepared according to the followingprocedure. To a solution of compound 1aa (0.27 g, 1 mmol) in carbontetrachloride, halogenation reagent (1 equiv) was added under N₂atmosphere. The reaction was allowed to stir at room temperature for2-2.5 h until TLC detected the reaction was complete (eluting withDCM:MeOH=10:1). After cooling to room temperature, filtering andconcentrating in vacuum, compound 1′″ was afforded by columnchromatography.

Table 5 provides exemplary conditions and compounds which were preparedaccording to the above general procedure.

TABLE 5 Exemplary Preparations of Compound 1′′′ Yield entry 1 reagent1′′′ % 45 1aa NBS

  1aab 34 46 1aa NCS

  1aac 25Compounds from Table 5 were analyzed by NMR and mass spectrometry. Datais provided below:1aab: ¹H NMR (400 MHz, DMSO-d₆) δ 12.15 (s, 1H), 8.12 (s, 1H), 7.48 (d,J=7.9 Hz, 1H), 7.18 (d, J=7.1 Hz, 1H), 7.04 (t, J=7.8 Hz, 1H), 4.33(ddd, J=6.8, 4.9, 1.6 Hz, 1H), 3.11 (dd, J=14.6, 4.9 Hz, 1H), 2.99 (dd,J=14.6, 7.2 Hz, 1H), 2.70 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 174.17,157.31, 133.52, 129.60, 121.54, 120.73, 117.72, 115.60, 112.70, 110.24,56.97, 27.77, 24.46; ESI-MS m/z 380.1 (M+Na)⁺; ESI-HRMS m/z calcd forC₁₃H₁₁BrClN₃NaO₂ (M+Na)⁺ 377.9616. found 377.9614.1aac: ¹H NMR (500 MHz, DMSO-d₆) δ 12.13 (s, 1H), 8.11 (s, 1H), 7.48 (d,J=7.9 Hz, 1H), 7.17 (dd, J=7.7, 0.8 Hz, 1H), 7.03 (t, J=7.8 Hz, 1H),4.32 (ddd, J=6.9, 4.9, 1.6 Hz, 1H), 3.11 (dd, J=14.7, 4.9 Hz, 1H), 2.99(dd, J=14.7, 7.3 Hz, 1H), 2.70 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ174.14, 157.25, 131.93, 129.33, 124.58, 121.60, 120.81, 117.86, 115.62,106.98, 56.91, 26.72, 24.43; ESI-MS m/z 310.0 (M−H)⁻; ESI-HRMS m/z calcdfor C₁₃H₁₀Cl₂N₃O₂ ⁻ (M−H)⁻ 310.0156. found 310.0168.

Example 7 Cellular Necrosis Inhibition Activity by Compounds ofStructure (I)

Compound (Ia) was prepared according to the general methods describedabove. Compound (Ia) and compound A (below) were screened foranti-necrotic activity utilizing human FADD-deficient Jurkat T cellschallenged with human TNF-alpha. FADD−/− Jurkat cells (Juo P, et al.Cell Growth Differ. 1999, 10(12):797-804) were seeded at the density of5*10⁵ cells/mL into 96 well white plates (Costar) at 100 μL/well.Compound A was identified as an inhibitor of necrosis in U.S. Pat. No.7,491,743, the full disclosure of which is incorporated herein byreference in its entirety. Cells were treated in duplicate with variousconcentrations of test compounds in the presence or absence of 10 ng/mlhuman TNF-alpha. (Cell Sciences). The compounds were also tested in thepresence of TNF-alpha and an Smac mimetic (10 nM of SM-164 (Lu et al,Cancer Res., 2008, 68:9384). Smac mimetics are a class of compounds thatinhibit IAPs and are thought to induce autocrine TNF-alpha productionand cell death. Accordingly, testing the compounds in the presence ofSmac provides further evidence of necrosis inhibitory activity.

After 30 hours, viability of the cells was determined using luminescentATP-based cell viability assay (CellTiter-Glo, Promega). Percentage ofprotection by the compound was calculated as a ratio of the cps (countsper second) value in the well treated with the test compound andTNF-alpha. to the cps value in the well treated with the compound alone.Assays were repeated twice, and the average EC₅₀ value over both assayswas calculated. Table 6 provides the average EC₅₀ (nM) values ofcompounds (Ia) and A for cell viability.

TABLE 6 Comparative EC₅₀ Data for Compounds (Ia) and 2 Compound EC50(nM) TNFα EC50 (nM) TNFα + Smac (Ia) 26.75 86.6 A 90.5 324

The data in Table 6 show that compound (Ia) is significantly more activethan compound A.

Example 8 Contribution of 6-Fluoro Substituent to Cellular NecrosisInhibition Activity

To investigate the contribution of the 6-fluoro substituent to thenecrosis inhibitory activity of compound (Ia), the activity of compound(Ia) was compared to the activity of the following structurally relatedcompounds (B-F below) according to the general procedures of Example 7,the results of which are summarized in Table 7:

TABLE 7 Comparative EC₅₀ Data for Compounds (Ia), B, C, D, E and FCompound EC50 (nM) TNFa EC50 (nM) TNFa + Smac (Ia) 26.75 86.6 B >100,000NA* C NA* NA* D NA* NA* E 2,330 8,330 F 10,500 >100,000 *NA = noactivity

The data in Table 7 show that the type of halogen substituent (Cl vs. F)and its position on the indole ring are both important contributors tothe increased activity of compound (Ia) relative to the comparativecompounds.

Example 9 RIP1 Kinase Inhibition by Compounds of Structure (I)

RIP1 kinase assays were performed using ADP-Glo assay (Promega)according to manufacturer's protocol. Reactions were performed in 50 mMHEPES, pH 7.5, 50 mM NaCl, 30 mM MgCl₂, 1 mM DTT, 0.05% bovine serumalbumin (BSA), 0.02% CHAPS buffer, containing 20 ng recombinantGST-human RIP1 kinase domain (amino acids 1-327), 50 mM ATP and 10serial dilutions of inhibitors. Recombinant GST-RIP1 was generated usingbaculoviral expression system in Sf9 cells. Protein was purified byglutathione affinity chromatography, followed by size exclusionchromatography. Reactions were performed for 4 hr at room temperatureand stopped by incubation with ADP-Glo reagent for 40 min at roomtemperature. Luminescent signal was developed by incubation with KinaseDetection reagent for 30 min at room temperature. Signal was determinedusing Victor3V platereader (Perkin Elmer). Non-linear regression tocalculate EC50 values was performed using Graph Pad Prism softwarepackage. Using these methods, Compound Ia was compared to Compound A forthe ability to inhibit RIP1 kinase enzymatic activity. As shown in FIG.1, the EC50 for inhibition of RIP1 kinase was 93.7 nM for Compound Ia(squares), substantially more potent than Compound A (circles), whichhad an EC50 of 255.9 nM.

Example 10 Cellular Necrosis Inhibition Activity of Compounds ofStructure (II)

Exemplary compounds of structure (II) were screened for anti-necroticactivity according to the general procedures of Example 7. Table 8provides EC₅₀ (nM) values for the tested compounds and a comparativecompound (C1) described in U.S. Patent Publication No. 2012/0122889, thefull disclosure of which is hereby incorporated by reference in itsentirety.

TABLE 8 Comparative EC₅₀ Data EC50 (nM) EC50 (nM) No. Structure TNFαTNFα + Smac II-1

159 11,000 II-2

 41   895 II-3

 89   426 II-4

220   867 II-5

235   892 II-6

 50   822 II-7

121   722 C1

343 Not tested

The data in Table 8 show that the tested compounds of structure (II) aresignificantly more active than compound C1.

Example 11 RIP1 Kinase Inhibition by Compounds of Structure (II)

RIP1 kinase assays were performed according to the general methodsdescribed in Example 9. Using these methods, compounds 1 am(5-((7-chloro-1H-indol-3-yl)methyl)-3-(3-fluorobenzyl)imidazolidine-2,4-dione)and 1ao(5-((7-chloro-1H-indol-3-yl)methyl)-3-(3,4-difluorobenzyl)imidazolidine-2,4-dione)were each found to have EC50 values of less than 100 nM.

Although various embodiments of the invention are disclosed herein, manyadaptations and modifications may be made within the scope of theinvention in accordance with the common general knowledge of thoseskilled in this art. Such modifications include the substitution ofknown equivalents for any aspect of the invention in order to achievethe same result in substantially the same way. Numeric ranges areinclusive of the numbers defining the range. The word “comprising” isused herein as an open-ended term, substantially equivalent to thephrase “including, but not limited to”, and the word “comprises” has acorresponding meaning. As used herein, the singular forms “a”, “an” and“the” include plural referents unless the context clearly dictatesotherwise. Thus, for example, reference to “a thing” includes more thanone such thing. Citation of references herein is not an admission thatsuch references are prior art to the present invention.

All of the U.S. patents, U.S. patent application publications, U.S.patent applications, foreign patents, foreign patent applications andnon-patent publications referred to in this specification or theattached Application Data Sheet are incorporated herein by reference, intheir entirety to the extent not inconsistent with the presentdescription.

The present application claims the benefit of the filing date of ChinesePatent Application No. 201410764426.4, filed Dec. 11, 2014, ChinesePatent Application No. 201410767595.3, filed Dec. 11, 2014, U.S.Provisional Patent Application Ser. No. 62/105,462, filed Jan. 20, 2015,entitled “POTENT INHIBITOR OF CELLULAR NECROSIS,” and U.S. ProvisionalPatent Application Ser. No. 62/105,475, filed Jan. 20, 2015, entitled“METHODS FOR PREPARATION OF CELLULAR NECROSIS INHIBITORS,” whichapplications are incorporated herein by reference, in their entirety.

From the foregoing it will be appreciated that, although specificembodiments of the invention have been described herein for purposes ofillustration, various modifications may be made without deviating fromthe spirit and scope of the invention. Accordingly, the invention is notlimited except as by the appended claims.

1. (canceled)
 2. A compound having the following structure (Ia):

or a pharmaceutically acceptable salt, prodrug or tautomer thereof.3-12. (canceled)
 13. A pharmaceutical composition comprising a compoundhaving the following structure (Ia):

or a pharmaceutically acceptable salt, prodrug or tautomer thereof, anda pharmaceutically acceptable carrier, diluent or excipient.
 14. Amethod for treating a necrotic cell disease, the method comprisingadministering an effective amount of a pharmaceutical composition to asubject in need thereof, wherein the pharmaceutical compositioncomprises a compound having the following structure (Ia):

or a pharmaceutically acceptable salt, prodrug or tautomer thereof, anda pharmaceutically acceptable carrier, diluent or excipient.
 15. Themethod of claim 14, wherein the necrotic cell disease is trauma,ischemia, stroke, cardiac infarction, infection, Gaucher's disease,Krabbe disease, sepsis, Parkinson's disease, Alzheimer's disease,amyotrophic lateral sclerosis, Huntington's disease, HIV-associateddementia, retinal degenerative disease, glaucoma, age-related maculardegeneration, rheumatoid arthritis, psoriasis, psoriatic arthritis orinflammatory bowel disease.
 16. A method for treating an inflammatorydisorder, the method comprising administering an effective amount of thecomposition of claim 13 to a subject in need thereof.
 17. The method ofclaim 16, wherein the inflammatory disorder is inflammatory boweldisease.
 18. The method of claim 16, wherein the inflammatory disorderis Crohn's disease or ulcerative colitis. 19-54. (canceled)
 55. Themethod of claim 15, wherein the necrotic cell disease is Parkinson'sdisease.
 56. The method of claim 15, wherein the necrotic cell diseaseis Alzheimer's disease.
 57. The method of claim 15, wherein the necroticcell disease is amyotrophic lateral sclerosis.
 58. The method of claim15, wherein the necrotic cell disease is Huntington's disease.
 59. Themethod of claim 15, wherein the necrotic cell disease is HIV-associateddementia.